Affinage

CRACR2A

EF-hand calcium-binding domain-containing protein 4B · UniProt Q9BSW2

Length
731 aa
Mass
83.2 kDa
Annotated
2026-06-09
15 papers in source corpus 10 papers cited in narrative 10 extracted findings
Cross-family judge vs UniProt: Affinage preferred faithfulness: 6/7 claims corpus-supported (86%)

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

CRACR2A (EFCAB4B) is a Ca2+-sensing protein, expressed as a short EF-hand isoform and a long large-Rab-GTPase isoform (Rab46), that couples calcium signaling to two distinct effector outputs: store-operated Ca2+ entry and dynein-based vesicle transport (PMID:20418871, PMID:30814157, PMID:31092558). The short isoform directly binds both Orai1 and STIM1 to form a Ca2+-dissociable ternary complex that stabilizes Orai1/STIM1 co-clustering and supports CRAC channel function; constitutive (Ca2+-insensitive) EF-hand mutants drive aberrant STIM1 clustering, Ca2+ overload, and cell death (PMID:20418871). In lymphocytes, a large isoform bearing EF-hand motifs, a proline-rich domain (PRD), and a prenylated Rab GTPase domain localizes to Golgi-proximal vesicles whose PRD–Vav1 interaction drives translocation to the immunological synapse, transmitting TCR signals through the Ca2+/NFAT and JNK/AP1 pathways to support Th1 and Th17 effector responses (PMID:27016526, PMID:29987160). Mechanistically, CRACR2A functions as a calcium-activated dynein adaptor: its coiled-coil domain assembles dynein and dynactin into a motile complex and its EF-hand domain confers Ca2+-dependent activation of motility, linking Ca2+ rises to retrograde endocytic transport (PMID:30814157). In endothelial cells the Rab46 isoform integrates histamine-driven G protein and Ca2+ signals to selectively route a P-selectin-negative Weibel-Palade body subset to the MTOC via dynein, with EF-hand Ca2+ binding triggering subsequent dispersal and selective cargo release [PMID:31092558, PMID:bio_10.1101_2025.03.03.641167]. In neutrophils CRACR2A associates with STIM1 after agonist stimulation to drive Ca2+ mobilization and β2 integrin activation, promoting recruitment (PMID:39601147). Biallelic loss-of-function mutations in CRACR2A cause late-onset combined immunodeficiency, and allele-specific reconstitution separates its SOCE function from its JNK-activating function (PMID:34908525).

Mechanistic history

Synthesis pass · year-by-year structured walk · 10 steps
  1. 2010 High

    Established CRACR2A as a Ca2+-sensing regulator of CRAC channels by showing it bridges Orai1 and STIM1 in a complex that disassembles when Ca2+ rises.

    Evidence Affinity purification, reciprocal Co-IP, siRNA knockdown, and EF-hand mutagenesis with confocal cluster imaging

    PMID:20418871

    Open questions at the time
    • Structural basis of the ternary complex not resolved
    • Did not address the existence of a second large isoform
  2. 2014 Medium

    Revealed that the CRACR2A locus produces a long endothelial isoform (Rab46) with an additional Rab GTPase domain that is functionally distinct from the CRAC-channel-regulating short form.

    Evidence cDNA cloning, Western blot, siRNA knockdown, and endothelial tube formation assay

    PMID:25475730

    Open questions at the time
    • Mechanism of contribution to tube formation unresolved
    • GTPase domain function not yet characterized
  3. 2016 High

    Defined the lymphocyte large isoform as a vesicle-associated Rab GTPase that uses a PRD-Vav1 interaction to deliver TCR signaling to the immunological synapse, positioning CRACR2A in NFAT and JNK pathways.

    Evidence Knockout mice, GTP-binding/prenylation mutants, CRACR2A-PRD/Vav1 Co-IP, live-cell vesicle imaging, and signaling assays

    PMID:27016526

    Open questions at the time
    • Identity of vesicle cargo at the synapse not defined
    • How Vav1 GEF activity feeds back on CRACR2A unclear
  4. 2018 High

    Demonstrated the physiological consequence of CRACR2A loss in T cells, linking it to Th1/Th17 effector function and in vivo immune responses.

    Evidence T cell-specific conditional knockout mice in LCMV and EAE models with Ca2+/NFAT and JNK readouts

    PMID:29987160

    Open questions at the time
    • Does not separate the SOCE versus JNK contributions to disease phenotypes
  5. 2019 High

    Established the core molecular mechanism: CRACR2A is a calcium-activated dynein adaptor whose coiled-coil assembles dynein-dynactin and whose EF-hand confers Ca2+-dependent motility, mediating endocytic transport.

    Evidence In vitro dynein motility reconstitution plus Ca2+ manipulation and live-cell puncta imaging in Jurkat T cells

    PMID:30814157

    Open questions at the time
    • Cargo of the endocytic transport process not identified
    • Single lab
  6. 2019 High

    Showed that the Rab46 isoform performs selective organelle sorting in endothelium, using dynein for MTOC retrograde transport and EF-hand Ca2+ sensing to disperse a specific WPB subset.

    Evidence Nucleotide-locked Rab46 mutants, dynein inhibition, Ca2+ imaging, and selective cargo release assays

    PMID:31092558

    Open questions at the time
    • Source of the dispersal-triggering Ca2+ not yet defined
    • Single lab
  7. 2021 Medium

    Identified the GTP-bound Rab46 effector proteome, confirming dynein and adding Na+/K+ ATPase subunit ATP1α1 as direct partners.

    Evidence Affinity purification of constitutively active vs inactive GFP-Rab46 with LC-MS/MS and biochemical/imaging validation

    PMID:33603063

    Open questions at the time
    • Functional role of the ATP1α1 interaction not pursued
    • No mechanistic follow-up
  8. 2021 High

    Proved CRACR2A is disease-relevant in humans and genetically dissected its two signaling functions via allele-specific reconstitution.

    Evidence Patient allele characterization and CRACR2A-KO T cell reconstitution with SOCE, JNK, and cytokine readouts

    PMID:34908525

    Open questions at the time
    • Structural basis for the E278D SOCE-specific defect not defined
    • Spectrum of clinical phenotypes incompletely mapped
  9. 2024 High

    Extended CRACR2A function to innate immunity, showing a STIM1-coupled Ca2+ mobilization role that activates β2 integrin and drives neutrophil recruitment, and provided a peptide inhibitor.

    Evidence Myeloid-specific KO mice, CRISPR KO in dHL-60 cells, rescue, CRACR2A-STIM1 Co-IP, Ca2+ and integrin assays, intravital microscopy, and a coiled-coil blocking peptide

    PMID:39601147

    Open questions at the time
    • Whether neutrophil function uses the short or large isoform not fully resolved
    • Relationship to dynein-adaptor activity in neutrophils unaddressed
  10. 2025 Medium

    Pinpointed TPC2 endolysosomal Ca2+ as the specific signal driving EF-hand-dependent Rab46 detachment from the MTOC to enable Ang2-positive WPB secretion.

    Evidence Ca2+ imaging, high-resolution microscopy, pharmacological TPC2 inhibition/activation, and Ang2 secretion assays (preprint)

    PMID:bio_10.1101_2025.03.03.641167

    Open questions at the time
    • No genetic confirmation of TPC2 specificity
    • Single lab, preprint

Open questions

Synthesis pass · forward-looking unresolved questions
  • How the short CRAC-regulatory isoform and the large dynein-adaptor/Rab isoform are differentially deployed across cell types, and whether their functions intersect within a single cell, remains unresolved.
  • No unified structural model linking EF-hand Ca2+ sensing, coiled-coil dynein assembly, and Rab GTPase activity
  • Isoform-specific upstream regulation not mapped

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0003924 GTPase activity 3 GO:0140299 molecular sensor activity 3 GO:0060090 molecular adaptor activity 2 GO:0098772 molecular function regulator activity 2 GO:0008092 cytoskeletal protein binding 1
Localization
GO:0005815 microtubule organizing center 2 GO:0005886 plasma membrane 2 GO:0031410 cytoplasmic vesicle 2 GO:0005794 Golgi apparatus 1
Pathway
R-HSA-168256 Immune System 4 R-HSA-162582 Signal Transduction 3 R-HSA-5653656 Vesicle-mediated transport 2 R-HSA-9609507 Protein localization 2
Partners
ATP1A1ORAI1STIM1VAV1
Complex memberships
Orai1-STIM1-CRACR2A ternary CRAC complexdynein-dynactin motile complex

Evidence

Reading pass · 10 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2010 CRACR2A (EFCAB4B) directly interacts with both Orai1 and STIM1, forming a ternary complex that dissociates at elevated Ca2+ concentrations. CRACR2A is required for co-clustering of Orai1 and STIM1 upon store depletion, and an EF-hand mutant of CRACR2A that cannot sense Ca2+ constitutively enhanced STIM1 clustering, elevated cytoplasmic Ca2+, and induced cell death. Affinity protein purification, siRNA knockdown, EF-hand mutagenesis, co-immunoprecipitation, confocal imaging of Orai1/STIM1 clusters Nature cell biology High 20418871
2016 The CRACR2A locus encodes a lymphocyte-specific large Rab GTPase isoform containing EF-hand motifs, a proline-rich domain (PRD), and a Rab GTPase domain with an unconventional prenylation site. This isoform localizes to vesicles near the Golgi (dependent on GTP binding and prenylation), which translocate to the immunological synapse upon TCR stimulation to activate Ca2+/NFAT and JNK signaling pathways. The interaction between the PRD of CRACR2A and the GEF Vav1 is required for accumulation of these vesicles at the immunological synapse. Gene silencing, knockout mice, domain mutagenesis (GTP-binding and prenylation mutants), co-immunoprecipitation of CRACR2A-PRD with Vav1, live-cell imaging of vesicle translocation, Ca2+ and JNK signaling assays Science signaling High 27016526
2019 CRACR2A acts as a calcium-activated dynein adaptor: its coiled-coil domain joins dynein and dynactin into a motile complex, and its EF-hand domain activates this motility in a calcium-dependent manner in vitro. In Jurkat T cells, elevation of intracellular calcium activates CRACR2A-mediated dynein transport, and TCR activation induces formation of CRACR2A puncta at the plasma membrane that associate with the actin cortex and then travel along microtubules in an endocytic process. In vitro dynein motility reconstitution assay, intracellular Ca2+ manipulation in Jurkat T cells, live-cell imaging of CRACR2A puncta, microtubule and actin co-localization The Journal of cell biology High 30814157
2019 Rab46 (CRACR2A long isoform) in endothelial cells integrates histamine (H1 receptor/G protein) and Ca2+ signals to regulate selective Weibel-Palade body (WPB) trafficking: GTP-bound Rab46 on a P-selectin-negative WPB subset mediates dynein-dependent retrograde transport to the microtubule organizing center (MTOC) upon histamine stimulation, and subsequently Ca2+ binding to Rab46's EF-hand triggers dispersal of MTOC-clustered WPBs, thereby diverting non-inflammatory cargo away from the plasma membrane. Rab46 localization by imaging, GTP/GDP-locked Rab46 mutants, dynein inhibition, intracellular Ca2+ measurement, selective cargo release assay (P-selectin vs. non-inflammatory cargo) The Journal of cell biology High 31092558
2014 Endothelial cells express a long variant of CRACR2A (CRACR2A-L / Rab46) that is approximately twice the molecular mass of the short form and contains an additional C-terminal Rab GTPase domain; siRNA against CRACR2A in endothelial cells depletes this long form but has no effect on CRAC channel function, and CRACR2A-L makes a positive contribution to endothelial tube formation. siRNA knockdown, Western blot, full-length cDNA cloning, sequence analysis, endothelial tube formation assay Biochemical and biophysical research communications Medium 25475730
2018 CRACR2A promotes Th1 differentiation and effector function of Th17 cells downstream of TCR signaling: conditional T cell-specific deletion of CRACR2A in mice reduced Th1 responses to LCMV infection and conferred resistance to EAE, with impaired Ca2+/NFAT and JNK/AP1 pathway activation due to deficient TCR signal transmission. Conditional knockout mice (T cell-specific Cre), viral infection model, EAE model, transcript analysis, Ca2+ and JNK signaling assays Journal of immunology High 29987160
2021 Biallelic loss-of-function mutations in CRACR2A cause late-onset combined immunodeficiency in humans. Patient T cells show reduced CRACR2A protein, dampened SOCE (Ca2+ entry), and reduced JNK phosphorylation. Reconstitution experiments in CRACR2A-deleted T cells showed that the E278D allele specifically impairs SOCE/cytokine production (not JNK), while the R144G/E300* truncation impairs both JNK phosphorylation and SOCE, establishing allele-specific separation of CRACR2A's two signaling functions. Patient allele characterization, CRACR2A KO T cell reconstitution with individual mutant alleles, SOCE measurement (Ca2+ imaging), JNK phosphorylation assay, cytokine production assay eLife High 34908525
2021 Affinity purification coupled to LC-MS/MS identified dynein and Na+/K+ ATPase subunit alpha 1 (ATP1α1) as validated binding partners (effectors) of GTP-bound Rab46 (CRACR2A) in endothelial cells, confirmed by biochemical and imaging approaches. Affinity purification with constitutively active vs. inactive GFP-Rab46, LC-MS/MS, biochemical validation, imaging Scientific reports Medium 33603063
2024 In neutrophils, CRACR2A rapidly associates with STIM1 after agonist stimulation and facilitates Ca2+ mobilization, which increases the ligand-binding function of β2 integrin and promotes neutrophil adhesion, crawling, and transmigration. A palmitoylated 20-mer peptide from the coiled-coil region of CRACR2A blocks STIM1-CRACR2A interaction, inhibiting Ca2+ mobilization and β2 integrin activation. Myeloid-specific Cracr2a conditional KO mice, CRISPR/Cas9 KO in dHL-60 cells, CRACR2A rescue overexpression, immunoprecipitation of CRACR2A-STIM1 complex, cytosolic Ca2+ mobilization assay, flow cytometry for integrin activation, intravital microscopy, blocking peptide experiments Circulation High 39601147
2025 Ca2+ released specifically from TPC2 (two-pore channel 2) endolysosomal channels is required for the EF-hand-dependent detachment of Rab46 (CRACR2A) from the MTOC, thereby enabling secretion of Ang2-positive Weibel-Palade bodies in endothelial cells. Pharmacological TPC2 inhibition increased Rab46 clustering at the MTOC and decreased Ang2 secretion, while a TPC2 agonist had the opposite effect. Ca2+ imaging, high-resolution light microscopy, pharmacological inhibition/activation of TPC2 (Ned19, tetrandrine, TPC2-A1-N), Rab46 localization at MTOC, Ang2 secretion assay bioRxivpreprint Medium bio_10.1101_2025.03.03.641167

Source papers

Stage 0 corpus · 15 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2010 A novel EF-hand protein, CRACR2A, is a cytosolic Ca2+ sensor that stabilizes CRAC channels in T cells. Nature cell biology 196 20418871
2019 CRACR2a is a calcium-activated dynein adaptor protein that regulates endocytic traffic. The Journal of cell biology 44 30814157
2016 A large Rab GTPase encoded by CRACR2A is a component of subsynaptic vesicles that transmit T cell activation signals. Science signaling 32 27016526
2019 Rab46 integrates Ca2+ and histamine signaling to regulate selective cargo release from Weibel-Palade bodies. The Journal of cell biology 26 31092558
2018 Vigorous Physical Activity Is Associated with Lower Risk of Metastatic-Lethal Progression in Prostate Cancer and Hypomethylation in the CRACR2A Gene. Cancer epidemiology, biomarkers & prevention : a publication of the American Association for Cancer Research, cosponsored by the American Society of Preventive Oncology 22 30464020
2018 CRACR2A-Mediated TCR Signaling Promotes Local Effector Th1 and Th17 Responses. Journal of immunology (Baltimore, Md. : 1950) 19 29987160
2014 Expression of a long variant of CRACR2A that belongs to the Rab GTPase protein family in endothelial cells. Biochemical and biophysical research communications 17 25475730
2021 Biallelic mutations in calcium release activated channel regulator 2A (CRACR2A) cause a primary immunodeficiency disorder. eLife 16 34908525
2018 A genome-wide association study identifies an association between variants in EFCAB4B gene and periodontal disease in an Italian isolated population. Journal of periodontal research 16 30284742
2024 Neutrophil CRACR2A Promotes Neutrophil Recruitment in Sterile Inflammation and Ischemic Stroke. Circulation 11 39601147
2021 Affinity-based proteomics reveals novel binding partners for Rab46 in endothelial cells. Scientific reports 7 33603063
2022 Cytogenomic Characterization of a Novel de novo Balanced Reciprocal Translocation t(1;12) by Genome Sequencing Leading to Fusion Gene Formation of EYA3/EFCAB4b. Molecular syndromology 3 36588754
2023 Rab46: a novel player in mast cell function. Discovery immunology 2 38567292
2025 RNA-seq analysis reveals transcriptome changes in livers from Efcab4b knockout mice. Biochemistry and biophysics reports 1 40034259
2022 CRACR2A associated immunodeficiency - the supporting factor takes center stage. Cell calcium 1 35151051

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