{"gene":"ACBD3","run_date":"2026-06-09T22:02:38","timeline":{"discoveries":[{"year":2001,"finding":"GCP60 (ACBD3) was identified as a peripheral Golgi membrane protein that interacts with the C-terminal cytoplasmic domain of the integral membrane protein giantin; overexpression of the GCP60 C-terminal domain caused Golgi disassembly and blocked ER-to-Golgi protein transport.","method":"Yeast two-hybrid screening, immunofluorescence, immunoelectron microscopy, overexpression assay","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — yeast two-hybrid plus subcellular localization with functional consequence (Golgi disruption), single lab with two orthogonal methods","pmids":["11590181"],"is_preprint":false},{"year":2001,"finding":"PAP7 (ACBD3) interacts with both the mitochondrial peripheral-type benzodiazepine receptor (PBR) and the cytosolic PKA regulatory subunit RIα; overexpression of full-length PAP7 increased hCG-induced steroid production, while a dominant-negative partial PAP7 and antisense oligonucleotides inhibited hormone-stimulated cholesterol transport and steroidogenesis in MA-10 Leydig cells.","method":"Yeast two-hybrid, GST pulldown, antisense oligonucleotide knockdown, steroid production assay","journal":"Molecular endocrinology (Baltimore, Md.)","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — GST pulldown plus functional loss-of-function assay, single lab with multiple orthogonal methods","pmids":["11731621"],"is_preprint":false},{"year":2003,"finding":"PAP7 (ACBD3) functions as an A-kinase anchoring protein (AKAP) that localizes to the trans-Golgi apparatus and mitochondria in Leydig cells; inhibition of PAP7 expression reduced hormone-induced cholesterol transport into mitochondria and decreased steroid formation, suggesting it targets PKA to PBR-rich organelles.","method":"Immunofluorescence confocal microscopy, antisense inhibition, steroid formation assay","journal":"The Journal of steroid biochemistry and molecular biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct localization experiment tied to functional consequence, loss-of-function with defined phenotype, single lab","pmids":["12943713"],"is_preprint":false},{"year":2006,"finding":"GCP60 (ACBD3) preferentially interacts with a caspase-generated golgin-160 fragment (residues 140–311) and prevents its nuclear translocation; cells overexpressing GCP60 showed increased sensitivity to staurosporine-induced apoptosis.","method":"Yeast two-hybrid, co-immunoprecipitation, overexpression/localization assay, apoptosis assay","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — multiple methods (yeast two-hybrid, co-IP, overexpression) in single lab demonstrating binding and functional consequence","pmids":["16870622"],"is_preprint":false},{"year":2007,"finding":"A single redox-sensitive cysteine (Cys-463) in GCP60 (ACBD3) is critical for its interaction with the golgin-160 caspase fragment (residues 140–311); mutation of Cys-463 abolished interaction in vitro and disrupted Golgi retention of the fragment; oxidation by H2O2 or a nitric oxide donor restored the interaction.","method":"Site-directed mutagenesis, in vitro binding assay, cellular localization assay, oxidation experiments","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 / Moderate — active-site mutagenesis with in vitro reconstitution and cellular validation, single lab but multiple orthogonal methods","pmids":["17711851"],"is_preprint":false},{"year":2007,"finding":"ACBD3 associates with the Golgi in neurons and interphase progenitor cells but becomes cytosolic upon Golgi fragmentation during mitosis; ACBD3 interacts with Numb through an essential Numb domain, and cytosolic ACBD3 acts synergistically with Numb to specify neural cell fates; loss- and gain-of-function mouse mutants share phenotypic similarities linking ACBD3 to asymmetric cell division.","method":"Co-immunoprecipitation, live/fixed-cell imaging, loss-of-function/gain-of-function mouse models","journal":"Cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal co-IP, direct subcellular localization tied to functional outcome, in vivo genetic models with defined phenotype","pmids":["17418793"],"is_preprint":false},{"year":2011,"finding":"ACBD3 interacts with multiple Aichi virus non-structural proteins (2B, 2BC, 2C, 3A, 3AB) and directly with PI4KB; this ACBD3–PI4KB interaction recruits PI4KB to viral RNA replication sites, enabling PI4P synthesis essential for Aichi virus RNA replication; knockdown of ACBD3 or PI4KB suppressed replication.","method":"Co-immunoprecipitation, siRNA knockdown, immunofluorescence microscopy, viral replication assay","journal":"The EMBO journal","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal co-IP, siRNA knockdown with defined replication phenotype, localization data; replicated across multiple subsequent studies","pmids":["22124328"],"is_preprint":false},{"year":2012,"finding":"Multiple picornavirus 3A proteins (Aichi virus, bovine kobuvirus, poliovirus, coxsackievirus B2/B3/B5, HRV14) co-purify with ACBD3; ACBD3 itself binds PI4KIIIβ in the absence of 3A; alanine-scanning mutagenesis of Aichi virus 3A identified residues that selectively abolish PI4KIIIβ co-purification without affecting ACBD3 binding; N-terminal glycines of some 3A proteins are myristoylated.","method":"Affinity purification with Strep-tag, mass spectrometry, Western blotting, alanine-scanning mutagenesis, siRNA knockdown, viral replication assay","journal":"Journal of virology","confidence":"High","confidence_rationale":"Tier 1 / Strong — affinity purification–MS with mutagenesis and functional validation, multiple viral systems tested, replicated key findings from PMID 22124328","pmids":["22258260"],"is_preprint":false},{"year":2012,"finding":"ACBD3 recruits the protein phosphatase PPM1L to ER–Golgi membrane contact sites via its GOLD domain, implicating ACBD3 in ceramide trafficking regulation at the ER–Golgi interface.","method":"Co-immunoprecipitation, domain mapping, subcellular fractionation/localization","journal":"FEBS letters","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — co-IP with domain mapping showing GOLD domain mediates interaction and recruits PPM1L to ER-Golgi contact sites, single lab","pmids":["22796112"],"is_preprint":false},{"year":2013,"finding":"ACBD3 interacts with TBC1D22A and TBC1D22B (putative Rab33 GAPs) via the same binding site on ACBD3 used by PI4KB; TBC1D22A/B and PI4KB interactions with ACBD3 are mutually exclusive, suggesting a competitive regulatory mechanism for PI4KB recruitment.","method":"Affinity purification–mass spectrometry, mammalian two-hybrid, co-immunoprecipitation, domain mapping","journal":"mBio","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — AP-MS plus two-hybrid with domain competition mapping, single lab with multiple orthogonal methods","pmids":["23572552"],"is_preprint":false},{"year":2013,"finding":"ACBD3 forms a complex with Rhes and mutant huntingtin (mHtt) in the striatum; ACBD3 levels are elevated in HD striatum; ACBD3 deletion abolishes mHtt-mediated neurotoxicity, while overexpression increases it, placing ACBD3 downstream of Rhes/mHtt as a mediator of HD cytotoxicity.","method":"Co-immunoprecipitation, ACBD3 deletion/overexpression, cell viability assay, Western blot in HD mouse brain and patient tissue","journal":"Cell reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP plus genetic loss/gain-of-function with defined neurotoxicity phenotype, single lab with multiple methods","pmids":["24012756"],"is_preprint":false},{"year":2013,"finding":"ACBD3 interacts with poliovirus 3A proteins at viral RNA replication sites; siRNA-mediated downregulation of ACBD3 significantly increased poliovirus replication, indicating ACBD3 can negatively modulate enterovirus replication; the amino acid at position 12 of 3A influences sensitivity to this effect.","method":"Co-immunoprecipitation, siRNA knockdown, viral growth assay, replicon assay, immunofluorescence","journal":"Journal of virology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP, siRNA KD with replication phenotype, mutagenesis, single lab","pmids":["23926333"],"is_preprint":false},{"year":2013,"finding":"ACBD3 depletion did not affect PI4KIIIβ recruitment to coxsackievirus B3 (CVB3) replication organelles and did not impair CVB3 RNA replication, demonstrating that CVB3 recruits PI4KIIIβ by an ACBD3-independent mechanism (NEGATIVE finding for CVB3).","method":"siRNA knockdown, immunofluorescence, viral replication assay","journal":"Journal of virology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — siRNA KD with clear negative functional readout, single lab with multiple methods; negative result for CVB3 specifically","pmids":["24352456"],"is_preprint":false},{"year":2014,"finding":"The viral protein/ACBD3/PI4KB complex stimulates PI4KB kinase activity in vitro; Aichi virus 3A and 3AB proteins stimulate PI4KB activity through forming a 3A(3AB)/ACBD3/PI4KB complex, enhancing PI4P synthesis at replication organelles and facilitating viral replication complex formation.","method":"In vitro kinase assay, siRNA knockdown, immunofluorescence, PI4P quantification","journal":"Journal of virology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro kinase assay demonstrating enzymatic activation, combined with siRNA and localization studies, single lab with multiple orthogonal methods","pmids":["24672044"],"is_preprint":false},{"year":2016,"finding":"NMR structure of the PI4KB–ACBD3 complex was determined; ACBD3 recruits PI4KB to membranes both in vitro and in vivo, and membrane recruitment of PI4KB by ACBD3 increases its enzymatic activity; ACBD3:PI4KB complex formation is essential for proper Golgi PI4P homeostasis.","method":"NMR structure determination, in vitro membrane recruitment assay, enzymatic activity assay, cellular localization experiments","journal":"Scientific reports","confidence":"High","confidence_rationale":"Tier 1 / Strong — NMR structure combined with in vitro reconstitution of membrane recruitment and enzymatic activation, validated in vivo; multiple orthogonal methods","pmids":["27009356"],"is_preprint":false},{"year":2016,"finding":"Salmonella effectors SseF and SseG interact directly with ACBD3; SseG binds ACBD3 alone, while SseF binding requires SseG; ACBD3 knockdown reduces Golgi association of Salmonella-containing vacuoles, and ACBD3-interaction-deficient SseF/SseG mutants display an intracellular replication defect.","method":"Yeast two-hybrid, co-immunoprecipitation, GST pulldown, siRNA knockdown, confocal microscopy, bacterial replication assay","journal":"mBio","confidence":"High","confidence_rationale":"Tier 2 / Strong — yeast two-hybrid, co-IP, GST pulldown, and siRNA knockdown with defined vacuole-positioning and replication phenotype; multiple orthogonal methods with mutagenesis","pmids":["27406559"],"is_preprint":false},{"year":2016,"finding":"Crystal structure of the ACBD3 GOLD domain revealed a unique N terminus that mediates interaction with Aichi virus 3A; hydrogen-deuterium exchange mass spectrometry mapped the PI4KIIIβ–ACBD3 and ACBD3–3A interfaces; 3A directly activates PI4KIIIβ and this is sensitized by ACBD3; rationally designed interface mutations abrogated kinase activation by ACBD3.","method":"Crystal structure determination, HDX-MS, in vitro kinase reconstitution, site-directed mutagenesis","journal":"Structure (London, England : 1993)","confidence":"High","confidence_rationale":"Tier 1 / Strong — crystal structure, HDX-MS interface mapping, in vitro kinase reconstitution, and mutagenesis in one study; multiple orthogonal Tier 1 methods","pmids":["27989622"],"is_preprint":false},{"year":2017,"finding":"Crystal structures of Aichi virus and bovine kobuvirus 3A proteins in complex with the ACBD3 GOLD domain showed that viral 3A proteins act as molecular harnesses to stabilize ACBD3 at target membranes; molecular dynamics simulation revealed 3A-mediated ACBD3 stabilization at lipid bilayers.","method":"Crystal structure determination, molecular dynamics simulation","journal":"Structure (London, England : 1993)","confidence":"High","confidence_rationale":"Tier 1 / Moderate — multiple crystal structures plus MD simulation with functional interpretation, single lab","pmids":["28065508"],"is_preprint":false},{"year":2017,"finding":"ACBD3 interacts with EV71 3A protein; this interaction is required for EV71 RNA replication and plaque formation; EV71 3A redirects ACBD3 to viral replication sites; I44A or H54Y substitutions in 3A disrupt ACBD3 binding and impair replication.","method":"cDNA library screening, co-immunoprecipitation, ACBD3 CRISPR knockout/knockdown, immunofluorescence, viral replication assay, mutagenesis","journal":"Scientific reports","confidence":"High","confidence_rationale":"Tier 2 / Strong — co-IP, CRISPR KO with rescue, mutagenesis, and replication phenotype; multiple orthogonal methods","pmids":["28303920"],"is_preprint":false},{"year":2017,"finding":"EV71 3A protein stimulates the ACBD3–PI4KB interaction; ACBD3 is required for PI4KB recruitment to EV71 RNA replication sites; EV71 infection induces PI4P production in an ACBD3- and PI4KB-dependent manner; I44A or H54Y in 3A abolish stimulation of ACBD3–PI4KB interaction.","method":"Co-immunoprecipitation, siRNA knockdown, immunofluorescence, PI4P quantification, mutagenesis","journal":"Journal of virology","confidence":"High","confidence_rationale":"Tier 2 / Strong — co-IP, siRNA KD with PI4P quantification and localization readouts, site-directed mutagenesis; multiple orthogonal methods","pmids":["28701404"],"is_preprint":false},{"year":2017,"finding":"ACBD3 interacts with Golgin45 via its GOLD domain; ACBD3 co-expression increases Golgin45 Golgi targeting; ACBD3 recruits TBC1D22 (a Rab33b GAP) to a multi-protein complex containing Golgin45 and GRASP55, suggesting a scaffolding role in organizing Golgi stacking proteins.","method":"Proteomics, co-immunoprecipitation, confocal microscopy, domain mapping","journal":"FEBS letters","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — co-IP and proteomics with subcellular localization data, single lab","pmids":["28777890"],"is_preprint":false},{"year":2018,"finding":"AiV non-structural proteins (2B, 2BC, 2C, 3A, 3AB) interact with ACBD3, OSBP, VAP-A/B, and SAC1; ACBD3 mediates recruitment of OSBP-VAP cholesterol transport machinery to AiV replication organelles through protein–protein interactions; silencing OSBP, VAP-A/B, or SAC1 inhibited AiV replication; cholesterol accumulates at AiV replication organelles in an OSBP-dependent manner.","method":"Co-immunoprecipitation, siRNA knockdown, immunofluorescence, cholesterol staining, electron microscopy","journal":"Journal of virology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP, siRNA KD with replication and lipid phenotypes, EM; single lab, multiple orthogonal methods","pmids":["29367253"],"is_preprint":false},{"year":2019,"finding":"ACBD3 knockout impaired replication of representative viruses from four enterovirus and two rhinovirus species; PI4KB recruitment to replication organelles requires ACBD3; absence of ACBD3 causes 3A mis-localization to ER instead of Golgi; ACB and CAR domains of ACBD3 are dispensable, while other domains are required for 3A-mediated PI4KB recruitment.","method":"CRISPR knockout, rescue with ACBD3/PI4KB mutants, immunofluorescence, viral replication assay, domain deletion analysis","journal":"mBio","confidence":"High","confidence_rationale":"Tier 2 / Strong — CRISPR KO with domain-specific rescue and multiple viral species; defined mechanistic epistasis; multiple orthogonal methods","pmids":["30755512"],"is_preprint":false},{"year":2019,"finding":"SAXS analysis showed that the ACBD3:PI4KB complex adopts highly flexible conformations (both compact and extended), while 14-3-3:PI4KB:Rab11 has 2:1:1 stoichiometry; membrane is required for formation of the ACBD3:PI4KB:Rab11 complex at physiological concentrations.","method":"Small-angle X-ray scattering (SAXS), in vitro reconstitution","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — SAXS and in vitro reconstitution demonstrating complex flexibility and membrane dependence, single lab","pmids":["30679637"],"is_preprint":false},{"year":2019,"finding":"Crystal structures of ACBD3 GOLD domain complexed with 3A proteins from poliovirus, EV-A71, EV-D68, and rhinovirus B14 revealed convergent structural mechanisms for 3A–ACBD3 interaction; 3A–3A interactions drive assembly of ACBD3–3A heterotetramers; structure-guided mutations disrupting these interfaces impaired PI4KB recruitment and enterovirus replication.","method":"Crystal structure determination, molecular dynamics, co-immunoprecipitation, viral replication assay, mutagenesis","journal":"PLoS pathogens","confidence":"High","confidence_rationale":"Tier 1 / Strong — multiple crystal structures with mutagenesis and functional replication assays across multiple enterovirus species","pmids":["31381608"],"is_preprint":false},{"year":2019,"finding":"ACBD3 is required for FAPP2-mediated glucosylceramide transport; ACBD3 knockdown causes Golgi fragmentation, FAPP2 dispersal from trans-Golgi network, and abnormal sphingolipid metabolism; re-expression of full-length ACBD3 rescues these defects.","method":"Co-immunoprecipitation, siRNA knockdown, quantitative lipidomics, confocal microscopy, rescue experiment","journal":"Journal of molecular cell biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP, siRNA KD with lipidomics and localization phenotype, rescue; single lab with multiple orthogonal methods","pmids":["29750412"],"is_preprint":false},{"year":2021,"finding":"ACBD3 knockout cells exhibit enlarged Golgi with absence of stacks and ribbon-like formation, confirming ACBD3 role in Golgi stacking; cholesterol levels and mitochondrial structure/function are not altered in ACBD3-KO HEK293 and HeLa cells; decreased sphingomyelins with normal ceramide and sphingomyelin synthase activity reveal ACBD3 role in ceramide transport from ER to Golgi.","method":"CRISPR knockout, electron microscopy, Golgi morphology analysis, lipidomics, mitochondrial function assays","journal":"International journal of molecular sciences","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — CRISPR KO with defined Golgi morphology and lipidomic phenotype; negative finding for mitochondrial function is informative; single lab","pmids":["34298889"],"is_preprint":false},{"year":2021,"finding":"ACBD3 directly interacts with KDEL receptor and recruits PKA to the Golgi; ACBD3 depletion causes accelerated retrograde trafficking of KDEL receptor by altering its interaction with PKA and Arf1/ArfGAP1, leading to increased Arf1-GTP-dependent tubular carrier formation; ACBD3 functions as a negative regulator of PKA activity on KDEL receptor.","method":"Proximity-based in vivo tagging, co-immunoprecipitation, siRNA knockdown, live-cell imaging, trafficking assay","journal":"BMC biology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — proximity labeling, co-IP, and siRNA KD with defined trafficking phenotype; single lab with multiple orthogonal methods","pmids":["34493279"],"is_preprint":false},{"year":2022,"finding":"The Golgi-resident ACBD3 recognizes and concentrates ligand-bound STING at specialized ER–Golgi contact sites (non-canonical ER exit sites); ACBD3 depletion impairs STING ER-to-Golgi trafficking and type-I interferon responses.","method":"Unbiased proteomic screen, super-resolution microscopy, live-cell imaging, ACBD3 siRNA knockdown, STING trafficking assay, IFN reporter assay","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 2 / Strong — unbiased proteomics, super-resolution microscopy, live-cell imaging, and siRNA KD with defined trafficking and signaling phenotypes; multiple orthogonal methods","pmids":["36543137"],"is_preprint":false},{"year":2023,"finding":"ACBD3 is recruited to the Golgi by two redundant mechanisms: (1) an MWT374-376 motif in the ACBD3 region upstream of the GOLD domain, which interacts with golgins golgin-45 and giantin; (2) interaction with SCFD1 (a Sec1/Munc-18 protein) and SEC22B (a SNARE); CRISPR-KO of SCFD1 causes ACBD3 to become cytosolic, acting upstream of golgin interactions.","method":"CRISPR knockout, unbiased proteomics, mutagenesis (MWT motif), co-immunoprecipitation, immunofluorescence","journal":"Molecular biology of the cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — CRISPR KO, unbiased proteomics, mutagenesis, and co-IP defining sequential Golgi recruitment mechanism; multiple orthogonal methods","pmids":["38134218"],"is_preprint":false},{"year":2023,"finding":"ACBD3 GOLD domain directly interacts with the regulatory subunit RII of PKA, recruiting PKA holoenzyme to the Golgi; forward trafficking of proteins from the ER triggers PKA activation (release of catalytic subunit from RII) at the Golgi; ACBD3 depletion reduces Golgi-localized RII and causes constitutive PKA activation and KDEL receptor retrograde transport.","method":"Co-immunoprecipitation, domain mapping, PKA activity assay, siRNA knockdown, KDEL receptor trafficking assay","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — co-IP with domain mapping, PKA activity assay, and functional trafficking readout; single lab with multiple orthogonal methods","pmids":["37044218"],"is_preprint":false},{"year":2024,"finding":"ACBD3 knockdown increases labile iron levels by promoting ferritinophagy, leading to ferroptosis sensitivity; this is coupled with reduced GPX4 levels and elevated polyunsaturated fatty acid-containing glycerophospholipids; knockdown of NCOA4 or Bafilomycin A1 treatment blocked ferritinophagy and impeded ferroptosis in ACBD3-depleted cells.","method":"siRNA knockdown, iron measurement, lipid peroxidation assay, lipidomics, ferritinophagy inhibition","journal":"Cell biology international","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — KD with multiple biochemical readouts and epistasis via ferritinophagy inhibitor; single lab","pmids":["38953242"],"is_preprint":false},{"year":2025,"finding":"ACBD3 colocalizes with TBEV NS4B at ER–Golgi contact sites and promotes TBEV infection; ACBD3 depletion inhibits virus replication and causes abnormal ER transformation and reduced virion release; the proviral mechanism is independent of PI4KB recruitment, requiring the full-length ACBD3 to coordinate ER-Golgi coupling.","method":"siRNA knockdown, proximity proteomics, confocal/electron microscopy, viral replication assay, virion release assay","journal":"Journal of virology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — proximity screen, siRNA KD with defined replication and morphological phenotype; negative finding for PI4KB mechanism; single lab","pmids":["40207930"],"is_preprint":false},{"year":2025,"finding":"ACBD3 promotes primary lung cancer growth by recruiting PI4KB to the Golgi, enhancing oncogenic secretion in chromosome 1q-amplified cells; conversely, in chromosome 1q-diploid cells, ACBD3 suppresses metastasis by inhibiting NOTCH signaling and reducing cell motility.","method":"CRISPR knockout, co-immunoprecipitation, cell migration/invasion assay, xenograft model, NOTCH signaling assay","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — CRISPR KO with in vitro and in vivo phenotypes and defined pathway (PI4KB/NOTCH); single lab, context-dependent findings","pmids":["40189704"],"is_preprint":false}],"current_model":"ACBD3 is a multifunctional Golgi-resident scaffolding protein that (1) directly recruits PI4KIIIβ (PI4KB) to Golgi/TGN membranes via its GOLD domain, increasing PI4KB enzymatic activity and local PI4P production essential for Golgi homeostasis and hijacked by picornaviruses for replication organelle biogenesis; (2) acts as an A-kinase anchoring protein (AKAP) at the Golgi and mitochondria by binding PKA-RIα/RII, coordinating cAMP-dependent steroidogenesis and KDEL receptor trafficking; (3) maintains Golgi stack integrity—required for FAPP2-mediated glucosylceramide transport and ceramide/sphingolipid homeostasis; (4) is itself recruited to the Golgi by a two-step mechanism involving SCFD1/SEC22B (upstream SNARE pathway) and golgins giantin/golgin-45 acting on an MWT motif in its pre-GOLD region; (5) concentrates ligand-activated STING at ER–Golgi contact sites to drive ER export and innate immune signaling; (6) interacts with Numb during mitosis-induced Golgi fragmentation to regulate asymmetric neural cell-fate specification; and (7) modulates apoptotic signaling through redox-sensitive (Cys-463) interaction with caspase-generated golgin-160 fragments."},"narrative":{"mechanistic_narrative":"ACBD3 (GCP60/PAP7) is a peripheral Golgi-membrane scaffolding protein that organizes lipid-modifying and signaling machinery on Golgi/TGN membranes and at ER–Golgi contact sites [PMID:11590181, PMID:27009356]. Its central scaffolding output is the direct recruitment of the lipid kinase PI4KB to membranes through its GOLD domain, an interaction defined structurally by NMR that both anchors PI4KB to the Golgi and stimulates its enzymatic activity to maintain Golgi PI4P homeostasis [PMID:27009356, PMID:27989622]. ACBD3 is itself targeted to the Golgi by a two-step mechanism: the Sec1/Munc-18 protein SCFD1 with the SNARE SEC22B acts upstream of an MWT374-376 motif that binds the golgins giantin and golgin-45 [PMID:38134218, PMID:11590181, PMID:28777890]. Beyond PI4KB, ACBD3 acts as an A-kinase anchoring protein, binding PKA regulatory subunits (RIα and RII via the GOLD domain) to position PKA at the Golgi and at mitochondria, where it couples cholesterol transport to hormone-stimulated steroidogenesis and where it controls cargo-triggered PKA activation governing KDEL-receptor retrograde trafficking [PMID:11731621, PMID:12943713, PMID:37044218, PMID:34493279]. ACBD3 is required for Golgi stack integrity and for FAPP2-mediated glucosylceramide transport and ER-to-Golgi ceramide/sphingolipid flux, with knockout producing enlarged, unstacked Golgi and altered sphingolipid pools [PMID:29750412, PMID:34298889]. The same membrane-coupling activity is extensively exploited by pathogens: picornavirus 3A proteins clamp ACBD3 onto replication-organelle membranes to recruit and activate PI4KB for viral PI4P synthesis, and the OSBP–VAP cholesterol-transport machinery is co-opted through ACBD3 as well [PMID:22124328, PMID:22258260, PMID:27989622, PMID:28065508, PMID:31381608, PMID:29367253]. ACBD3 also concentrates ligand-activated STING at ER–Golgi contact sites to drive ER export and type-I interferon responses [PMID:36543137], partners with Numb during mitotic Golgi fragmentation to influence asymmetric neural cell-fate specification [PMID:17418793], and modulates apoptotic signaling through a redox-sensitive Cys-463 interaction with a caspase-generated golgin-160 fragment [PMID:17711851]. Salmonella effectors SseF/SseG bind ACBD3 to position Salmonella-containing vacuoles at the Golgi [PMID:27406559], underscoring ACBD3 as a recurrently hijacked membrane-organizing hub.","teleology":[{"year":2001,"claim":"Established ACBD3 as a Golgi-associated protein whose integrity is functionally required for Golgi architecture and ER-to-Golgi transport, defining its core cellular compartment.","evidence":"Yeast two-hybrid against giantin C-terminus with immuno-EM and overexpression-induced Golgi disassembly","pmids":["11590181"],"confidence":"Medium","gaps":["Mechanism of how giantin binding stabilizes Golgi structure unresolved","Did not identify downstream effectors recruited by ACBD3","Endogenous loss-of-function not tested"]},{"year":2001,"claim":"Revealed a second functional pool of ACBD3 as a PKA/PBR-associated factor at mitochondria controlling steroidogenesis, framing it as an AKAP coupling cAMP signaling to cholesterol transport.","evidence":"Yeast two-hybrid and GST pulldown with RIα/PBR plus antisense knockdown and steroid assays in MA-10 Leydig cells","pmids":["11731621","12943713"],"confidence":"Medium","gaps":["Direct demonstration of PKA targeting to mitochondria not structurally defined","Relationship between Golgi and mitochondrial pools unclear","Later work found no mitochondrial structural/functional defect in KO cells"]},{"year":2007,"claim":"Connected ACBD3 to apoptotic signaling via a redox-gated interaction, showing a single cysteine acts as an oxidation sensor controlling sequestration of a pro-apoptotic golgin-160 fragment.","evidence":"Site-directed mutagenesis of Cys-463 with in vitro binding, cellular localization, and H2O2/NO oxidation experiments","pmids":["17711851","16870622"],"confidence":"High","gaps":["Physiological signals that oxidize Cys-463 in vivo not defined","Quantitative contribution to apoptosis regulation unknown","Structural basis of redox switch not solved"]},{"year":2007,"claim":"Demonstrated that cell-cycle-dependent relocalization of ACBD3 (Golgi to cytosol upon mitotic Golgi fragmentation) couples organelle dynamics to a developmental cell-fate decision via Numb.","evidence":"Reciprocal co-IP, live/fixed imaging, and loss/gain-of-function mouse models","pmids":["17418793"],"confidence":"High","gaps":["Molecular mechanism linking cytosolic ACBD3 to Numb-dependent fate output unclear","Whether scaffolding of PKA/PI4KB participates not addressed"]},{"year":2011,"claim":"Identified ACBD3 as the direct bridge recruiting PI4KB to picornavirus replication sites, establishing the ACBD3–PI4KB axis as the molecular basis for viral PI4P-dependent replication-organelle biogenesis.","evidence":"Co-IP of ACBD3 with Aichi virus non-structural proteins and PI4KB plus siRNA knockdown and viral replication assays","pmids":["22124328","22258260"],"confidence":"High","gaps":["Whether ACBD3–PI4KB binding alters kinase activity not yet shown at this stage","Cellular (non-viral) PI4KB recruitment role inferred but untested here"]},{"year":2013,"claim":"Refined the PI4KB-recruitment model as competitive and context-dependent, showing TBC1D22 GAPs occupy the same ACBD3 site as PI4KB and that ACBD3 can negatively modulate some enteroviruses.","evidence":"AP-MS, mammalian two-hybrid, domain-competition mapping, and siRNA knockdown with viral growth assays","pmids":["23572552","23926333","24012756"],"confidence":"Medium","gaps":["Regulatory logic switching ACBD3 between PI4KB and TBC1D22 binding unknown","Mechanism of negative modulation of poliovirus replication unresolved","Rhes/mHtt complex role in neurodegeneration mechanistically thin"]},{"year":2014,"claim":"Showed the viral 3A/ACBD3/PI4KB ternary complex directly stimulates PI4KB catalytic activity, moving ACBD3 from a passive tether to an allosteric activator of lipid kinase output.","evidence":"In vitro kinase assay with reconstituted complexes plus siRNA and PI4P quantification","pmids":["24672044"],"confidence":"High","gaps":["Quantitative contribution of ACBD3 versus 3A to activation not fully separated","Membrane requirement not yet incorporated"]},{"year":2016,"claim":"Provided the structural basis for ACBD3 function: NMR and crystal structures defined the PI4KB and 3A interfaces on the GOLD domain and proved membrane recruitment of PI4KB by ACBD3 raises its activity and sets Golgi PI4P levels.","evidence":"NMR and crystal structure determination, HDX-MS interface mapping, in vitro membrane recruitment and kinase reconstitution with interface mutagenesis","pmids":["27009356","27989622"],"confidence":"High","gaps":["Full-length ACBD3 architecture not resolved","How upstream golgin/SCFD1 anchoring positions the GOLD domain unknown"]},{"year":2017,"claim":"Defined ACBD3 as a recurrently hijacked membrane hub across diverse pathogens, with viral 3A proteins acting as molecular harnesses stabilizing ACBD3 at membranes and bacterial SseF/SseG positioning Salmonella vacuoles via ACBD3.","evidence":"Crystal structures with MD simulation, plus yeast two-hybrid, co-IP, GST pulldown, and bacterial/viral replication assays with interface mutagenesis","pmids":["28065508","27406559","28303920","28701404"],"confidence":"High","gaps":["Endogenous host signals that mimic 3A harnessing not identified","Whether bacterial recruitment involves PI4KB/PKA scaffolds unclear"]},{"year":2017,"claim":"Extended the scaffolding model to Golgi structural proteins, showing ACBD3 GOLD-domain interactions organize golgin-45, GRASP55, and TBC1D22 into a stacking-related complex.","evidence":"Proteomics, co-IP, domain mapping, and confocal microscopy","pmids":["28777890"],"confidence":"Medium","gaps":["Direct role of this complex in stack formation not functionally proven here","Competition with PI4KB for the same site not reconciled in vivo"]},{"year":2019,"claim":"Established ACBD3 as broadly essential for enterovirus/rhinovirus replication through domain-specific PI4KB recruitment, and clarified its native lipid-transport role in glucosylceramide and ceramide trafficking and Golgi stacking.","evidence":"CRISPR knockout with domain-specific rescue, lipidomics, FAPP2 localization, and SAXS conformational analysis of the ACBD3:PI4KB complex","pmids":["30755512","29750412","30679637","31381608","29367253"],"confidence":"High","gaps":["Mechanism coupling ACBD3 to FAPP2/ceramide transport molecularly undefined","How conformational flexibility of ACBD3:PI4KB is regulated on membranes unclear","CVB3 uses an ACBD3-independent route, leaving virus-specificity determinants open"]},{"year":2021,"claim":"Resolved a native AKAP function at the Golgi, showing ACBD3 tethers PKA to the KDEL receptor and acts as a negative regulator of PKA-driven retrograde trafficking and Arf1-dependent tubular carriers.","evidence":"Proximity-based tagging, co-IP, siRNA knockdown, and live-cell trafficking assays; CRISPR-KO Golgi morphology and lipidomics","pmids":["34493279","34298889"],"confidence":"Medium","gaps":["Signal triggering PKA release at the Golgi not defined here","Reconciliation of negative regulation with mitochondrial steroidogenic role incomplete"]},{"year":2022,"claim":"Identified ACBD3 as an innate-immune trafficking factor that concentrates ligand-bound STING at non-canonical ER–Golgi contact sites to drive ER export and type-I interferon responses.","evidence":"Unbiased proteomics, super-resolution and live-cell imaging, siRNA knockdown, STING trafficking and IFN reporter assays","pmids":["36543137"],"confidence":"High","gaps":["Whether STING concentration uses PI4KB/PKA scaffolds or a distinct mode unknown","Structural basis of STING recognition not defined"]},{"year":2023,"claim":"Defined the upstream logic of ACBD3 membrane targeting and confirmed a GOLD-domain PKA-RII interaction coupling forward cargo flux to Golgi PKA activation.","evidence":"CRISPR knockout of SCFD1, MWT-motif mutagenesis, unbiased proteomics, co-IP, PKA activity and KDEL trafficking assays","pmids":["38134218","37044218"],"confidence":"High","gaps":["How SCFD1/SEC22B SNARE machinery hands ACBD3 to golgins mechanistically unresolved","Cargo sensor linking secretion to PKA activation unidentified"]},{"year":2025,"claim":"Broadened ACBD3's roles to ferroptosis control, PI4KB-independent flavivirus support, and context-dependent oncogenic versus metastasis-suppressive functions in lung cancer.","evidence":"siRNA/CRISPR knockdown with iron/lipid-peroxidation measurements, proximity proteomics and viral assays for TBEV, and xenograft/migration assays with NOTCH readouts","pmids":["38953242","40207930","40189704"],"confidence":"Medium","gaps":["Mechanism linking ACBD3 loss to NCOA4-dependent ferritinophagy unclear","PI4KB-independent ER–Golgi coupling activity for TBEV undefined","Determinants of pro- versus anti-tumor switch by 1q copy number unresolved"]},{"year":null,"claim":"How a single Golgi scaffold partitions its competing GOLD-domain interactions (PI4KB, PKA-RII, golgin-45, TBC1D22, viral 3A, STING) into distinct spatial and temporal outputs, and what signals govern this switching, remains unresolved.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No integrated regulatory model coordinating mutually exclusive partners","Full-length structure on native membranes lacking","In vivo physiological hierarchy among ACBD3's roles undefined"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0060090","term_label":"molecular adaptor activity","supporting_discovery_ids":[14,16,29,20,27]},{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[14,13,30]},{"term_id":"GO:0008289","term_label":"lipid binding","supporting_discovery_ids":[25,26]}],"localization":[{"term_id":"GO:0005794","term_label":"Golgi apparatus","supporting_discovery_ids":[0,2,14,26,29]},{"term_id":"GO:0005783","term_label":"endoplasmic reticulum","supporting_discovery_ids":[8,28,32]},{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[5]},{"term_id":"GO:0005739","term_label":"mitochondrion","supporting_discovery_ids":[1,2]}],"pathway":[{"term_id":"R-HSA-5653656","term_label":"Vesicle-mediated transport","supporting_discovery_ids":[0,25,27]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[6,7,22,24,15,28]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[28]},{"term_id":"R-HSA-1430728","term_label":"Metabolism","supporting_discovery_ids":[25,26,1]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[30,27,1]}],"complexes":["ACBD3:PI4KB complex","ACBD3:PI4KB:Rab11 membrane complex","golgin-45:GRASP55:TBC1D22 stacking complex","ACBD3:PKA holoenzyme (AKAP) complex"],"partners":["PI4KB","GIANTIN","GOLGA1/GOLGIN-45","PRKAR1A","TBC1D22A","SCFD1","SEC22B","KDELR"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q9H3P7","full_name":"Golgi resident protein GCP60","aliases":["Acyl-CoA-binding domain-containing protein 3","Golgi complex-associated protein 1","GOCAP1","Golgi phosphoprotein 1","GOLPH1","PBR- and PKA-associated protein 7","Peripheral benzodiazepine receptor-associated protein PAP7"],"length_aa":528,"mass_kda":60.6,"function":"Involved in the maintenance of Golgi structure by interacting with giantin, affecting protein transport between the endoplasmic reticulum and Golgi (PubMed:11590181). Involved in hormone-induced steroid biosynthesis in testicular Leydig cells (By similarity). Recruits PI4KB to the Golgi apparatus membrane; enhances the enzyme activity of PI4KB activity via its membrane recruitment thereby increasing the local concentration of the substrate in the vicinity of the kinase (PubMed:27009356) (Microbial infection) Plays an essential role in Aichi virus RNA replication by recruiting PI4KB at the viral replication sites","subcellular_location":"Golgi apparatus membrane; Mitochondrion","url":"https://www.uniprot.org/uniprotkb/Q9H3P7/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/ACBD3","classification":"Not Classified","n_dependent_lines":1,"n_total_lines":1208,"dependency_fraction":0.0008278145695364238},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"SEC61B","stoichiometry":4.0}],"url":"https://opencell.sf.czbiohub.org/search/ACBD3","total_profiled":1310},"omim":[{"mim_id":"620867","title":"ARMADILLO-LIKE HELICAL DOMAIN-CONTAINING PROTEIN 3; ARMH3","url":"https://www.omim.org/entry/620867"},{"mim_id":"616880","title":"TBC1 DOMAIN FAMILY, MEMBER 22B; TBC1D22B","url":"https://www.omim.org/entry/616880"},{"mim_id":"616879","title":"TBC1 DOMAIN FAMILY, MEMBER 22A; TBC1D22A","url":"https://www.omim.org/entry/616879"},{"mim_id":"606809","title":"ACYL-CoA-BINDING DOMAIN-CONTAINING PROTEIN 3; ACBD3","url":"https://www.omim.org/entry/606809"},{"mim_id":"602758","title":"PHOSPHATIDYLINOSITOL 4-KINASE, BETA; PI4KB","url":"https://www.omim.org/entry/602758"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Supported","locations":[{"location":"Golgi apparatus","reliability":"Supported"}],"tissue_specificity":"Low tissue specificity","tissue_distribution":"Detected in all","driving_tissues":[],"url":"https://www.proteinatlas.org/search/ACBD3"},"hgnc":{"alias_symbol":["GCP60","PAP7"],"prev_symbol":["GOLPH1","GOCAP1"]},"alphafold":{"accession":"Q9H3P7","domains":[{"cath_id":"1.20.80.10","chopping":"68-168","consensus_level":"medium","plddt":91.7714,"start":68,"end":168},{"cath_id":"2.60.120.680","chopping":"378-452_472-526","consensus_level":"high","plddt":92.2493,"start":378,"end":526}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9H3P7","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q9H3P7-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q9H3P7-F1-predicted_aligned_error_v6.png","plddt_mean":78.75},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=ACBD3","jax_strain_url":"https://www.jax.org/strain/search?query=ACBD3"},"sequence":{"accession":"Q9H3P7","fasta_url":"https://rest.uniprot.org/uniprotkb/Q9H3P7.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q9H3P7/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q9H3P7"}},"corpus_meta":[{"pmid":"22124328","id":"PMC_22124328","title":"ACBD3-mediated 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metabolism","date":"2024-11-15","source":"bioRxiv","url":"https://doi.org/10.1101/2024.11.15.623757","citation_count":0,"is_preprint":true}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":27419,"output_tokens":8368,"usd":0.103888,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":18212,"output_tokens":5848,"usd":0.11863,"stage2_stop_reason":"end_turn"},"total_usd":0.222518,"stage1_batch_id":"msgbatch_0179Z2pxRYBsXi2xADneafRw","stage2_batch_id":"msgbatch_01LaY9ZaVdgjWArcZo9eqQE5","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2001,\n      \"finding\": \"GCP60 (ACBD3) was identified as a peripheral Golgi membrane protein that interacts with the C-terminal cytoplasmic domain of the integral membrane protein giantin; overexpression of the GCP60 C-terminal domain caused Golgi disassembly and blocked ER-to-Golgi protein transport.\",\n      \"method\": \"Yeast two-hybrid screening, immunofluorescence, immunoelectron microscopy, overexpression assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — yeast two-hybrid plus subcellular localization with functional consequence (Golgi disruption), single lab with two orthogonal methods\",\n      \"pmids\": [\"11590181\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2001,\n      \"finding\": \"PAP7 (ACBD3) interacts with both the mitochondrial peripheral-type benzodiazepine receptor (PBR) and the cytosolic PKA regulatory subunit RIα; overexpression of full-length PAP7 increased hCG-induced steroid production, while a dominant-negative partial PAP7 and antisense oligonucleotides inhibited hormone-stimulated cholesterol transport and steroidogenesis in MA-10 Leydig cells.\",\n      \"method\": \"Yeast two-hybrid, GST pulldown, antisense oligonucleotide knockdown, steroid production assay\",\n      \"journal\": \"Molecular endocrinology (Baltimore, Md.)\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — GST pulldown plus functional loss-of-function assay, single lab with multiple orthogonal methods\",\n      \"pmids\": [\"11731621\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"PAP7 (ACBD3) functions as an A-kinase anchoring protein (AKAP) that localizes to the trans-Golgi apparatus and mitochondria in Leydig cells; inhibition of PAP7 expression reduced hormone-induced cholesterol transport into mitochondria and decreased steroid formation, suggesting it targets PKA to PBR-rich organelles.\",\n      \"method\": \"Immunofluorescence confocal microscopy, antisense inhibition, steroid formation assay\",\n      \"journal\": \"The Journal of steroid biochemistry and molecular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct localization experiment tied to functional consequence, loss-of-function with defined phenotype, single lab\",\n      \"pmids\": [\"12943713\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"GCP60 (ACBD3) preferentially interacts with a caspase-generated golgin-160 fragment (residues 140–311) and prevents its nuclear translocation; cells overexpressing GCP60 showed increased sensitivity to staurosporine-induced apoptosis.\",\n      \"method\": \"Yeast two-hybrid, co-immunoprecipitation, overexpression/localization assay, apoptosis assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — multiple methods (yeast two-hybrid, co-IP, overexpression) in single lab demonstrating binding and functional consequence\",\n      \"pmids\": [\"16870622\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"A single redox-sensitive cysteine (Cys-463) in GCP60 (ACBD3) is critical for its interaction with the golgin-160 caspase fragment (residues 140–311); mutation of Cys-463 abolished interaction in vitro and disrupted Golgi retention of the fragment; oxidation by H2O2 or a nitric oxide donor restored the interaction.\",\n      \"method\": \"Site-directed mutagenesis, in vitro binding assay, cellular localization assay, oxidation experiments\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — active-site mutagenesis with in vitro reconstitution and cellular validation, single lab but multiple orthogonal methods\",\n      \"pmids\": [\"17711851\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"ACBD3 associates with the Golgi in neurons and interphase progenitor cells but becomes cytosolic upon Golgi fragmentation during mitosis; ACBD3 interacts with Numb through an essential Numb domain, and cytosolic ACBD3 acts synergistically with Numb to specify neural cell fates; loss- and gain-of-function mouse mutants share phenotypic similarities linking ACBD3 to asymmetric cell division.\",\n      \"method\": \"Co-immunoprecipitation, live/fixed-cell imaging, loss-of-function/gain-of-function mouse models\",\n      \"journal\": \"Cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal co-IP, direct subcellular localization tied to functional outcome, in vivo genetic models with defined phenotype\",\n      \"pmids\": [\"17418793\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"ACBD3 interacts with multiple Aichi virus non-structural proteins (2B, 2BC, 2C, 3A, 3AB) and directly with PI4KB; this ACBD3–PI4KB interaction recruits PI4KB to viral RNA replication sites, enabling PI4P synthesis essential for Aichi virus RNA replication; knockdown of ACBD3 or PI4KB suppressed replication.\",\n      \"method\": \"Co-immunoprecipitation, siRNA knockdown, immunofluorescence microscopy, viral replication assay\",\n      \"journal\": \"The EMBO journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal co-IP, siRNA knockdown with defined replication phenotype, localization data; replicated across multiple subsequent studies\",\n      \"pmids\": [\"22124328\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Multiple picornavirus 3A proteins (Aichi virus, bovine kobuvirus, poliovirus, coxsackievirus B2/B3/B5, HRV14) co-purify with ACBD3; ACBD3 itself binds PI4KIIIβ in the absence of 3A; alanine-scanning mutagenesis of Aichi virus 3A identified residues that selectively abolish PI4KIIIβ co-purification without affecting ACBD3 binding; N-terminal glycines of some 3A proteins are myristoylated.\",\n      \"method\": \"Affinity purification with Strep-tag, mass spectrometry, Western blotting, alanine-scanning mutagenesis, siRNA knockdown, viral replication assay\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — affinity purification–MS with mutagenesis and functional validation, multiple viral systems tested, replicated key findings from PMID 22124328\",\n      \"pmids\": [\"22258260\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"ACBD3 recruits the protein phosphatase PPM1L to ER–Golgi membrane contact sites via its GOLD domain, implicating ACBD3 in ceramide trafficking regulation at the ER–Golgi interface.\",\n      \"method\": \"Co-immunoprecipitation, domain mapping, subcellular fractionation/localization\",\n      \"journal\": \"FEBS letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — co-IP with domain mapping showing GOLD domain mediates interaction and recruits PPM1L to ER-Golgi contact sites, single lab\",\n      \"pmids\": [\"22796112\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"ACBD3 interacts with TBC1D22A and TBC1D22B (putative Rab33 GAPs) via the same binding site on ACBD3 used by PI4KB; TBC1D22A/B and PI4KB interactions with ACBD3 are mutually exclusive, suggesting a competitive regulatory mechanism for PI4KB recruitment.\",\n      \"method\": \"Affinity purification–mass spectrometry, mammalian two-hybrid, co-immunoprecipitation, domain mapping\",\n      \"journal\": \"mBio\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — AP-MS plus two-hybrid with domain competition mapping, single lab with multiple orthogonal methods\",\n      \"pmids\": [\"23572552\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"ACBD3 forms a complex with Rhes and mutant huntingtin (mHtt) in the striatum; ACBD3 levels are elevated in HD striatum; ACBD3 deletion abolishes mHtt-mediated neurotoxicity, while overexpression increases it, placing ACBD3 downstream of Rhes/mHtt as a mediator of HD cytotoxicity.\",\n      \"method\": \"Co-immunoprecipitation, ACBD3 deletion/overexpression, cell viability assay, Western blot in HD mouse brain and patient tissue\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP plus genetic loss/gain-of-function with defined neurotoxicity phenotype, single lab with multiple methods\",\n      \"pmids\": [\"24012756\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"ACBD3 interacts with poliovirus 3A proteins at viral RNA replication sites; siRNA-mediated downregulation of ACBD3 significantly increased poliovirus replication, indicating ACBD3 can negatively modulate enterovirus replication; the amino acid at position 12 of 3A influences sensitivity to this effect.\",\n      \"method\": \"Co-immunoprecipitation, siRNA knockdown, viral growth assay, replicon assay, immunofluorescence\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP, siRNA KD with replication phenotype, mutagenesis, single lab\",\n      \"pmids\": [\"23926333\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"ACBD3 depletion did not affect PI4KIIIβ recruitment to coxsackievirus B3 (CVB3) replication organelles and did not impair CVB3 RNA replication, demonstrating that CVB3 recruits PI4KIIIβ by an ACBD3-independent mechanism (NEGATIVE finding for CVB3).\",\n      \"method\": \"siRNA knockdown, immunofluorescence, viral replication assay\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — siRNA KD with clear negative functional readout, single lab with multiple methods; negative result for CVB3 specifically\",\n      \"pmids\": [\"24352456\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"The viral protein/ACBD3/PI4KB complex stimulates PI4KB kinase activity in vitro; Aichi virus 3A and 3AB proteins stimulate PI4KB activity through forming a 3A(3AB)/ACBD3/PI4KB complex, enhancing PI4P synthesis at replication organelles and facilitating viral replication complex formation.\",\n      \"method\": \"In vitro kinase assay, siRNA knockdown, immunofluorescence, PI4P quantification\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro kinase assay demonstrating enzymatic activation, combined with siRNA and localization studies, single lab with multiple orthogonal methods\",\n      \"pmids\": [\"24672044\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"NMR structure of the PI4KB–ACBD3 complex was determined; ACBD3 recruits PI4KB to membranes both in vitro and in vivo, and membrane recruitment of PI4KB by ACBD3 increases its enzymatic activity; ACBD3:PI4KB complex formation is essential for proper Golgi PI4P homeostasis.\",\n      \"method\": \"NMR structure determination, in vitro membrane recruitment assay, enzymatic activity assay, cellular localization experiments\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — NMR structure combined with in vitro reconstitution of membrane recruitment and enzymatic activation, validated in vivo; multiple orthogonal methods\",\n      \"pmids\": [\"27009356\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Salmonella effectors SseF and SseG interact directly with ACBD3; SseG binds ACBD3 alone, while SseF binding requires SseG; ACBD3 knockdown reduces Golgi association of Salmonella-containing vacuoles, and ACBD3-interaction-deficient SseF/SseG mutants display an intracellular replication defect.\",\n      \"method\": \"Yeast two-hybrid, co-immunoprecipitation, GST pulldown, siRNA knockdown, confocal microscopy, bacterial replication assay\",\n      \"journal\": \"mBio\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — yeast two-hybrid, co-IP, GST pulldown, and siRNA knockdown with defined vacuole-positioning and replication phenotype; multiple orthogonal methods with mutagenesis\",\n      \"pmids\": [\"27406559\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Crystal structure of the ACBD3 GOLD domain revealed a unique N terminus that mediates interaction with Aichi virus 3A; hydrogen-deuterium exchange mass spectrometry mapped the PI4KIIIβ–ACBD3 and ACBD3–3A interfaces; 3A directly activates PI4KIIIβ and this is sensitized by ACBD3; rationally designed interface mutations abrogated kinase activation by ACBD3.\",\n      \"method\": \"Crystal structure determination, HDX-MS, in vitro kinase reconstitution, site-directed mutagenesis\",\n      \"journal\": \"Structure (London, England : 1993)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — crystal structure, HDX-MS interface mapping, in vitro kinase reconstitution, and mutagenesis in one study; multiple orthogonal Tier 1 methods\",\n      \"pmids\": [\"27989622\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"Crystal structures of Aichi virus and bovine kobuvirus 3A proteins in complex with the ACBD3 GOLD domain showed that viral 3A proteins act as molecular harnesses to stabilize ACBD3 at target membranes; molecular dynamics simulation revealed 3A-mediated ACBD3 stabilization at lipid bilayers.\",\n      \"method\": \"Crystal structure determination, molecular dynamics simulation\",\n      \"journal\": \"Structure (London, England : 1993)\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — multiple crystal structures plus MD simulation with functional interpretation, single lab\",\n      \"pmids\": [\"28065508\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"ACBD3 interacts with EV71 3A protein; this interaction is required for EV71 RNA replication and plaque formation; EV71 3A redirects ACBD3 to viral replication sites; I44A or H54Y substitutions in 3A disrupt ACBD3 binding and impair replication.\",\n      \"method\": \"cDNA library screening, co-immunoprecipitation, ACBD3 CRISPR knockout/knockdown, immunofluorescence, viral replication assay, mutagenesis\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — co-IP, CRISPR KO with rescue, mutagenesis, and replication phenotype; multiple orthogonal methods\",\n      \"pmids\": [\"28303920\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"EV71 3A protein stimulates the ACBD3–PI4KB interaction; ACBD3 is required for PI4KB recruitment to EV71 RNA replication sites; EV71 infection induces PI4P production in an ACBD3- and PI4KB-dependent manner; I44A or H54Y in 3A abolish stimulation of ACBD3–PI4KB interaction.\",\n      \"method\": \"Co-immunoprecipitation, siRNA knockdown, immunofluorescence, PI4P quantification, mutagenesis\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — co-IP, siRNA KD with PI4P quantification and localization readouts, site-directed mutagenesis; multiple orthogonal methods\",\n      \"pmids\": [\"28701404\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"ACBD3 interacts with Golgin45 via its GOLD domain; ACBD3 co-expression increases Golgin45 Golgi targeting; ACBD3 recruits TBC1D22 (a Rab33b GAP) to a multi-protein complex containing Golgin45 and GRASP55, suggesting a scaffolding role in organizing Golgi stacking proteins.\",\n      \"method\": \"Proteomics, co-immunoprecipitation, confocal microscopy, domain mapping\",\n      \"journal\": \"FEBS letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — co-IP and proteomics with subcellular localization data, single lab\",\n      \"pmids\": [\"28777890\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"AiV non-structural proteins (2B, 2BC, 2C, 3A, 3AB) interact with ACBD3, OSBP, VAP-A/B, and SAC1; ACBD3 mediates recruitment of OSBP-VAP cholesterol transport machinery to AiV replication organelles through protein–protein interactions; silencing OSBP, VAP-A/B, or SAC1 inhibited AiV replication; cholesterol accumulates at AiV replication organelles in an OSBP-dependent manner.\",\n      \"method\": \"Co-immunoprecipitation, siRNA knockdown, immunofluorescence, cholesterol staining, electron microscopy\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP, siRNA KD with replication and lipid phenotypes, EM; single lab, multiple orthogonal methods\",\n      \"pmids\": [\"29367253\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"ACBD3 knockout impaired replication of representative viruses from four enterovirus and two rhinovirus species; PI4KB recruitment to replication organelles requires ACBD3; absence of ACBD3 causes 3A mis-localization to ER instead of Golgi; ACB and CAR domains of ACBD3 are dispensable, while other domains are required for 3A-mediated PI4KB recruitment.\",\n      \"method\": \"CRISPR knockout, rescue with ACBD3/PI4KB mutants, immunofluorescence, viral replication assay, domain deletion analysis\",\n      \"journal\": \"mBio\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — CRISPR KO with domain-specific rescue and multiple viral species; defined mechanistic epistasis; multiple orthogonal methods\",\n      \"pmids\": [\"30755512\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"SAXS analysis showed that the ACBD3:PI4KB complex adopts highly flexible conformations (both compact and extended), while 14-3-3:PI4KB:Rab11 has 2:1:1 stoichiometry; membrane is required for formation of the ACBD3:PI4KB:Rab11 complex at physiological concentrations.\",\n      \"method\": \"Small-angle X-ray scattering (SAXS), in vitro reconstitution\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — SAXS and in vitro reconstitution demonstrating complex flexibility and membrane dependence, single lab\",\n      \"pmids\": [\"30679637\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Crystal structures of ACBD3 GOLD domain complexed with 3A proteins from poliovirus, EV-A71, EV-D68, and rhinovirus B14 revealed convergent structural mechanisms for 3A–ACBD3 interaction; 3A–3A interactions drive assembly of ACBD3–3A heterotetramers; structure-guided mutations disrupting these interfaces impaired PI4KB recruitment and enterovirus replication.\",\n      \"method\": \"Crystal structure determination, molecular dynamics, co-immunoprecipitation, viral replication assay, mutagenesis\",\n      \"journal\": \"PLoS pathogens\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — multiple crystal structures with mutagenesis and functional replication assays across multiple enterovirus species\",\n      \"pmids\": [\"31381608\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"ACBD3 is required for FAPP2-mediated glucosylceramide transport; ACBD3 knockdown causes Golgi fragmentation, FAPP2 dispersal from trans-Golgi network, and abnormal sphingolipid metabolism; re-expression of full-length ACBD3 rescues these defects.\",\n      \"method\": \"Co-immunoprecipitation, siRNA knockdown, quantitative lipidomics, confocal microscopy, rescue experiment\",\n      \"journal\": \"Journal of molecular cell biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP, siRNA KD with lipidomics and localization phenotype, rescue; single lab with multiple orthogonal methods\",\n      \"pmids\": [\"29750412\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"ACBD3 knockout cells exhibit enlarged Golgi with absence of stacks and ribbon-like formation, confirming ACBD3 role in Golgi stacking; cholesterol levels and mitochondrial structure/function are not altered in ACBD3-KO HEK293 and HeLa cells; decreased sphingomyelins with normal ceramide and sphingomyelin synthase activity reveal ACBD3 role in ceramide transport from ER to Golgi.\",\n      \"method\": \"CRISPR knockout, electron microscopy, Golgi morphology analysis, lipidomics, mitochondrial function assays\",\n      \"journal\": \"International journal of molecular sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — CRISPR KO with defined Golgi morphology and lipidomic phenotype; negative finding for mitochondrial function is informative; single lab\",\n      \"pmids\": [\"34298889\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"ACBD3 directly interacts with KDEL receptor and recruits PKA to the Golgi; ACBD3 depletion causes accelerated retrograde trafficking of KDEL receptor by altering its interaction with PKA and Arf1/ArfGAP1, leading to increased Arf1-GTP-dependent tubular carrier formation; ACBD3 functions as a negative regulator of PKA activity on KDEL receptor.\",\n      \"method\": \"Proximity-based in vivo tagging, co-immunoprecipitation, siRNA knockdown, live-cell imaging, trafficking assay\",\n      \"journal\": \"BMC biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — proximity labeling, co-IP, and siRNA KD with defined trafficking phenotype; single lab with multiple orthogonal methods\",\n      \"pmids\": [\"34493279\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"The Golgi-resident ACBD3 recognizes and concentrates ligand-bound STING at specialized ER–Golgi contact sites (non-canonical ER exit sites); ACBD3 depletion impairs STING ER-to-Golgi trafficking and type-I interferon responses.\",\n      \"method\": \"Unbiased proteomic screen, super-resolution microscopy, live-cell imaging, ACBD3 siRNA knockdown, STING trafficking assay, IFN reporter assay\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — unbiased proteomics, super-resolution microscopy, live-cell imaging, and siRNA KD with defined trafficking and signaling phenotypes; multiple orthogonal methods\",\n      \"pmids\": [\"36543137\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"ACBD3 is recruited to the Golgi by two redundant mechanisms: (1) an MWT374-376 motif in the ACBD3 region upstream of the GOLD domain, which interacts with golgins golgin-45 and giantin; (2) interaction with SCFD1 (a Sec1/Munc-18 protein) and SEC22B (a SNARE); CRISPR-KO of SCFD1 causes ACBD3 to become cytosolic, acting upstream of golgin interactions.\",\n      \"method\": \"CRISPR knockout, unbiased proteomics, mutagenesis (MWT motif), co-immunoprecipitation, immunofluorescence\",\n      \"journal\": \"Molecular biology of the cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — CRISPR KO, unbiased proteomics, mutagenesis, and co-IP defining sequential Golgi recruitment mechanism; multiple orthogonal methods\",\n      \"pmids\": [\"38134218\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"ACBD3 GOLD domain directly interacts with the regulatory subunit RII of PKA, recruiting PKA holoenzyme to the Golgi; forward trafficking of proteins from the ER triggers PKA activation (release of catalytic subunit from RII) at the Golgi; ACBD3 depletion reduces Golgi-localized RII and causes constitutive PKA activation and KDEL receptor retrograde transport.\",\n      \"method\": \"Co-immunoprecipitation, domain mapping, PKA activity assay, siRNA knockdown, KDEL receptor trafficking assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — co-IP with domain mapping, PKA activity assay, and functional trafficking readout; single lab with multiple orthogonal methods\",\n      \"pmids\": [\"37044218\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"ACBD3 knockdown increases labile iron levels by promoting ferritinophagy, leading to ferroptosis sensitivity; this is coupled with reduced GPX4 levels and elevated polyunsaturated fatty acid-containing glycerophospholipids; knockdown of NCOA4 or Bafilomycin A1 treatment blocked ferritinophagy and impeded ferroptosis in ACBD3-depleted cells.\",\n      \"method\": \"siRNA knockdown, iron measurement, lipid peroxidation assay, lipidomics, ferritinophagy inhibition\",\n      \"journal\": \"Cell biology international\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — KD with multiple biochemical readouts and epistasis via ferritinophagy inhibitor; single lab\",\n      \"pmids\": [\"38953242\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"ACBD3 colocalizes with TBEV NS4B at ER–Golgi contact sites and promotes TBEV infection; ACBD3 depletion inhibits virus replication and causes abnormal ER transformation and reduced virion release; the proviral mechanism is independent of PI4KB recruitment, requiring the full-length ACBD3 to coordinate ER-Golgi coupling.\",\n      \"method\": \"siRNA knockdown, proximity proteomics, confocal/electron microscopy, viral replication assay, virion release assay\",\n      \"journal\": \"Journal of virology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — proximity screen, siRNA KD with defined replication and morphological phenotype; negative finding for PI4KB mechanism; single lab\",\n      \"pmids\": [\"40207930\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"ACBD3 promotes primary lung cancer growth by recruiting PI4KB to the Golgi, enhancing oncogenic secretion in chromosome 1q-amplified cells; conversely, in chromosome 1q-diploid cells, ACBD3 suppresses metastasis by inhibiting NOTCH signaling and reducing cell motility.\",\n      \"method\": \"CRISPR knockout, co-immunoprecipitation, cell migration/invasion assay, xenograft model, NOTCH signaling assay\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — CRISPR KO with in vitro and in vivo phenotypes and defined pathway (PI4KB/NOTCH); single lab, context-dependent findings\",\n      \"pmids\": [\"40189704\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"ACBD3 is a multifunctional Golgi-resident scaffolding protein that (1) directly recruits PI4KIIIβ (PI4KB) to Golgi/TGN membranes via its GOLD domain, increasing PI4KB enzymatic activity and local PI4P production essential for Golgi homeostasis and hijacked by picornaviruses for replication organelle biogenesis; (2) acts as an A-kinase anchoring protein (AKAP) at the Golgi and mitochondria by binding PKA-RIα/RII, coordinating cAMP-dependent steroidogenesis and KDEL receptor trafficking; (3) maintains Golgi stack integrity—required for FAPP2-mediated glucosylceramide transport and ceramide/sphingolipid homeostasis; (4) is itself recruited to the Golgi by a two-step mechanism involving SCFD1/SEC22B (upstream SNARE pathway) and golgins giantin/golgin-45 acting on an MWT motif in its pre-GOLD region; (5) concentrates ligand-activated STING at ER–Golgi contact sites to drive ER export and innate immune signaling; (6) interacts with Numb during mitosis-induced Golgi fragmentation to regulate asymmetric neural cell-fate specification; and (7) modulates apoptotic signaling through redox-sensitive (Cys-463) interaction with caspase-generated golgin-160 fragments.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"ACBD3 (GCP60/PAP7) is a peripheral Golgi-membrane scaffolding protein that organizes lipid-modifying and signaling machinery on Golgi/TGN membranes and at ER–Golgi contact sites [#0, #14]. Its central scaffolding output is the direct recruitment of the lipid kinase PI4KB to membranes through its GOLD domain, an interaction defined structurally by NMR that both anchors PI4KB to the Golgi and stimulates its enzymatic activity to maintain Golgi PI4P homeostasis [#14, #16]. ACBD3 is itself targeted to the Golgi by a two-step mechanism: the Sec1/Munc-18 protein SCFD1 with the SNARE SEC22B acts upstream of an MWT374-376 motif that binds the golgins giantin and golgin-45 [#29, #0, #20]. Beyond PI4KB, ACBD3 acts as an A-kinase anchoring protein, binding PKA regulatory subunits (RIα and RII via the GOLD domain) to position PKA at the Golgi and at mitochondria, where it couples cholesterol transport to hormone-stimulated steroidogenesis and where it controls cargo-triggered PKA activation governing KDEL-receptor retrograde trafficking [#1, #2, #30, #27]. ACBD3 is required for Golgi stack integrity and for FAPP2-mediated glucosylceramide transport and ER-to-Golgi ceramide/sphingolipid flux, with knockout producing enlarged, unstacked Golgi and altered sphingolipid pools [#25, #26]. The same membrane-coupling activity is extensively exploited by pathogens: picornavirus 3A proteins clamp ACBD3 onto replication-organelle membranes to recruit and activate PI4KB for viral PI4P synthesis, and the OSBP–VAP cholesterol-transport machinery is co-opted through ACBD3 as well [#6, #7, #16, #17, #24, #21]. ACBD3 also concentrates ligand-activated STING at ER–Golgi contact sites to drive ER export and type-I interferon responses [#28], partners with Numb during mitotic Golgi fragmentation to influence asymmetric neural cell-fate specification [#5], and modulates apoptotic signaling through a redox-sensitive Cys-463 interaction with a caspase-generated golgin-160 fragment [#4]. Salmonella effectors SseF/SseG bind ACBD3 to position Salmonella-containing vacuoles at the Golgi [#15], underscoring ACBD3 as a recurrently hijacked membrane-organizing hub.\",\n  \"teleology\": [\n    {\n      \"year\": 2001,\n      \"claim\": \"Established ACBD3 as a Golgi-associated protein whose integrity is functionally required for Golgi architecture and ER-to-Golgi transport, defining its core cellular compartment.\",\n      \"evidence\": \"Yeast two-hybrid against giantin C-terminus with immuno-EM and overexpression-induced Golgi disassembly\",\n      \"pmids\": [\"11590181\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism of how giantin binding stabilizes Golgi structure unresolved\", \"Did not identify downstream effectors recruited by ACBD3\", \"Endogenous loss-of-function not tested\"]\n    },\n    {\n      \"year\": 2001,\n      \"claim\": \"Revealed a second functional pool of ACBD3 as a PKA/PBR-associated factor at mitochondria controlling steroidogenesis, framing it as an AKAP coupling cAMP signaling to cholesterol transport.\",\n      \"evidence\": \"Yeast two-hybrid and GST pulldown with RIα/PBR plus antisense knockdown and steroid assays in MA-10 Leydig cells\",\n      \"pmids\": [\"11731621\", \"12943713\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct demonstration of PKA targeting to mitochondria not structurally defined\", \"Relationship between Golgi and mitochondrial pools unclear\", \"Later work found no mitochondrial structural/functional defect in KO cells\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Connected ACBD3 to apoptotic signaling via a redox-gated interaction, showing a single cysteine acts as an oxidation sensor controlling sequestration of a pro-apoptotic golgin-160 fragment.\",\n      \"evidence\": \"Site-directed mutagenesis of Cys-463 with in vitro binding, cellular localization, and H2O2/NO oxidation experiments\",\n      \"pmids\": [\"17711851\", \"16870622\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Physiological signals that oxidize Cys-463 in vivo not defined\", \"Quantitative contribution to apoptosis regulation unknown\", \"Structural basis of redox switch not solved\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Demonstrated that cell-cycle-dependent relocalization of ACBD3 (Golgi to cytosol upon mitotic Golgi fragmentation) couples organelle dynamics to a developmental cell-fate decision via Numb.\",\n      \"evidence\": \"Reciprocal co-IP, live/fixed imaging, and loss/gain-of-function mouse models\",\n      \"pmids\": [\"17418793\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular mechanism linking cytosolic ACBD3 to Numb-dependent fate output unclear\", \"Whether scaffolding of PKA/PI4KB participates not addressed\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Identified ACBD3 as the direct bridge recruiting PI4KB to picornavirus replication sites, establishing the ACBD3–PI4KB axis as the molecular basis for viral PI4P-dependent replication-organelle biogenesis.\",\n      \"evidence\": \"Co-IP of ACBD3 with Aichi virus non-structural proteins and PI4KB plus siRNA knockdown and viral replication assays\",\n      \"pmids\": [\"22124328\", \"22258260\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether ACBD3–PI4KB binding alters kinase activity not yet shown at this stage\", \"Cellular (non-viral) PI4KB recruitment role inferred but untested here\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Refined the PI4KB-recruitment model as competitive and context-dependent, showing TBC1D22 GAPs occupy the same ACBD3 site as PI4KB and that ACBD3 can negatively modulate some enteroviruses.\",\n      \"evidence\": \"AP-MS, mammalian two-hybrid, domain-competition mapping, and siRNA knockdown with viral growth assays\",\n      \"pmids\": [\"23572552\", \"23926333\", \"24012756\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Regulatory logic switching ACBD3 between PI4KB and TBC1D22 binding unknown\", \"Mechanism of negative modulation of poliovirus replication unresolved\", \"Rhes/mHtt complex role in neurodegeneration mechanistically thin\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Showed the viral 3A/ACBD3/PI4KB ternary complex directly stimulates PI4KB catalytic activity, moving ACBD3 from a passive tether to an allosteric activator of lipid kinase output.\",\n      \"evidence\": \"In vitro kinase assay with reconstituted complexes plus siRNA and PI4P quantification\",\n      \"pmids\": [\"24672044\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Quantitative contribution of ACBD3 versus 3A to activation not fully separated\", \"Membrane requirement not yet incorporated\"]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Provided the structural basis for ACBD3 function: NMR and crystal structures defined the PI4KB and 3A interfaces on the GOLD domain and proved membrane recruitment of PI4KB by ACBD3 raises its activity and sets Golgi PI4P levels.\",\n      \"evidence\": \"NMR and crystal structure determination, HDX-MS interface mapping, in vitro membrane recruitment and kinase reconstitution with interface mutagenesis\",\n      \"pmids\": [\"27009356\", \"27989622\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Full-length ACBD3 architecture not resolved\", \"How upstream golgin/SCFD1 anchoring positions the GOLD domain unknown\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Defined ACBD3 as a recurrently hijacked membrane hub across diverse pathogens, with viral 3A proteins acting as molecular harnesses stabilizing ACBD3 at membranes and bacterial SseF/SseG positioning Salmonella vacuoles via ACBD3.\",\n      \"evidence\": \"Crystal structures with MD simulation, plus yeast two-hybrid, co-IP, GST pulldown, and bacterial/viral replication assays with interface mutagenesis\",\n      \"pmids\": [\"28065508\", \"27406559\", \"28303920\", \"28701404\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Endogenous host signals that mimic 3A harnessing not identified\", \"Whether bacterial recruitment involves PI4KB/PKA scaffolds unclear\"]\n    },\n    {\n      \"year\": 2017,\n      \"claim\": \"Extended the scaffolding model to Golgi structural proteins, showing ACBD3 GOLD-domain interactions organize golgin-45, GRASP55, and TBC1D22 into a stacking-related complex.\",\n      \"evidence\": \"Proteomics, co-IP, domain mapping, and confocal microscopy\",\n      \"pmids\": [\"28777890\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct role of this complex in stack formation not functionally proven here\", \"Competition with PI4KB for the same site not reconciled in vivo\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Established ACBD3 as broadly essential for enterovirus/rhinovirus replication through domain-specific PI4KB recruitment, and clarified its native lipid-transport role in glucosylceramide and ceramide trafficking and Golgi stacking.\",\n      \"evidence\": \"CRISPR knockout with domain-specific rescue, lipidomics, FAPP2 localization, and SAXS conformational analysis of the ACBD3:PI4KB complex\",\n      \"pmids\": [\"30755512\", \"29750412\", \"30679637\", \"31381608\", \"29367253\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism coupling ACBD3 to FAPP2/ceramide transport molecularly undefined\", \"How conformational flexibility of ACBD3:PI4KB is regulated on membranes unclear\", \"CVB3 uses an ACBD3-independent route, leaving virus-specificity determinants open\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Resolved a native AKAP function at the Golgi, showing ACBD3 tethers PKA to the KDEL receptor and acts as a negative regulator of PKA-driven retrograde trafficking and Arf1-dependent tubular carriers.\",\n      \"evidence\": \"Proximity-based tagging, co-IP, siRNA knockdown, and live-cell trafficking assays; CRISPR-KO Golgi morphology and lipidomics\",\n      \"pmids\": [\"34493279\", \"34298889\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Signal triggering PKA release at the Golgi not defined here\", \"Reconciliation of negative regulation with mitochondrial steroidogenic role incomplete\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Identified ACBD3 as an innate-immune trafficking factor that concentrates ligand-bound STING at non-canonical ER–Golgi contact sites to drive ER export and type-I interferon responses.\",\n      \"evidence\": \"Unbiased proteomics, super-resolution and live-cell imaging, siRNA knockdown, STING trafficking and IFN reporter assays\",\n      \"pmids\": [\"36543137\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether STING concentration uses PI4KB/PKA scaffolds or a distinct mode unknown\", \"Structural basis of STING recognition not defined\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Defined the upstream logic of ACBD3 membrane targeting and confirmed a GOLD-domain PKA-RII interaction coupling forward cargo flux to Golgi PKA activation.\",\n      \"evidence\": \"CRISPR knockout of SCFD1, MWT-motif mutagenesis, unbiased proteomics, co-IP, PKA activity and KDEL trafficking assays\",\n      \"pmids\": [\"38134218\", \"37044218\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How SCFD1/SEC22B SNARE machinery hands ACBD3 to golgins mechanistically unresolved\", \"Cargo sensor linking secretion to PKA activation unidentified\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"Broadened ACBD3's roles to ferroptosis control, PI4KB-independent flavivirus support, and context-dependent oncogenic versus metastasis-suppressive functions in lung cancer.\",\n      \"evidence\": \"siRNA/CRISPR knockdown with iron/lipid-peroxidation measurements, proximity proteomics and viral assays for TBEV, and xenograft/migration assays with NOTCH readouts\",\n      \"pmids\": [\"38953242\", \"40207930\", \"40189704\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism linking ACBD3 loss to NCOA4-dependent ferritinophagy unclear\", \"PI4KB-independent ER–Golgi coupling activity for TBEV undefined\", \"Determinants of pro- versus anti-tumor switch by 1q copy number unresolved\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How a single Golgi scaffold partitions its competing GOLD-domain interactions (PI4KB, PKA-RII, golgin-45, TBC1D22, viral 3A, STING) into distinct spatial and temporal outputs, and what signals govern this switching, remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No integrated regulatory model coordinating mutually exclusive partners\", \"Full-length structure on native membranes lacking\", \"In vivo physiological hierarchy among ACBD3's roles undefined\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0060090\", \"supporting_discovery_ids\": [14, 16, 29, 20, 27]},\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [14, 13, 30]},\n      {\"term_id\": \"GO:0008289\", \"supporting_discovery_ids\": [25, 26]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005794\", \"supporting_discovery_ids\": [0, 2, 14, 26, 29]},\n      {\"term_id\": \"GO:0005783\", \"supporting_discovery_ids\": [8, 28, 32]},\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [5]},\n      {\"term_id\": \"GO:0005739\", \"supporting_discovery_ids\": [1, 2]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-5653656\", \"supporting_discovery_ids\": [0, 25, 27]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [6, 7, 22, 24, 15, 28]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [28]},\n      {\"term_id\": \"R-HSA-1430728\", \"supporting_discovery_ids\": [25, 26, 1]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [30, 27, 1]}\n    ],\n    \"complexes\": [\n      \"ACBD3:PI4KB complex\",\n      \"ACBD3:PI4KB:Rab11 membrane complex\",\n      \"golgin-45:GRASP55:TBC1D22 stacking complex\",\n      \"ACBD3:PKA holoenzyme (AKAP) complex\"\n    ],\n    \"partners\": [\n      \"PI4KB\",\n      \"giantin\",\n      \"GOLGA1/golgin-45\",\n      \"PRKAR1A\",\n      \"TBC1D22A\",\n      \"SCFD1\",\n      \"SEC22B\",\n      \"KDELR\"\n    ],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":8,"faith_total":8,"faith_pct":100.0}}