Affinage

ARL4A

ADP-ribosylation factor-like protein 4A · UniProt P40617

Length
200 aa
Mass
22.6 kDa
Annotated
2026-04-28
8 papers in source corpus 8 papers cited in narrative 8 extracted findings

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

ARL4A is a myristoylated small GTPase that acts as a compartment-specific recruitment platform at the plasma membrane, Golgi, and endosomes to coordinate cytoskeletal remodeling, membrane trafficking, and receptor sorting. At the plasma membrane, GTP-bound ARL4A recruits cytohesin Arf-GEFs via their PH domains to activate Arf6 signaling (PMID:17398095), and independently recruits the ELMO–DOCK180 complex through the ELMO Ras-binding domain to drive Rac-dependent actin remodeling (PMID:21930703); its stability and plasma membrane residence are regulated by Pak1-mediated phosphorylation at S143, which enables HYPK chaperone binding and protection from proteasomal degradation (PMID:35857868). At the Golgi, ARL4A engages the golgin GCC185 in a GTP-dependent manner to recruit CLASPs, maintain Golgi integrity, and support endosome-to-Golgi transport (PMID:22159419), while at endosomes it binds the ESCRT-II subunit VPS36 to stabilize ESCRT-III association, limit USP8-mediated deubiquitination, and thereby attenuate lysosomal degradation of EGFR (PMID:38030597).

Mechanistic history

Synthesis pass · year-by-year structured walk · 7 steps
  1. 2000 Medium

    Initial characterization established that ARL4A is N-terminally myristoylated and can localize to nuclei/nucleoli, interacting with importin-α via a C-terminal NLS, providing the first molecular handles for understanding its trafficking and regulation.

    Evidence Immunofluorescence, yeast two-hybrid, in vitro binding, and myristoylation assay in mammalian cells

    PMID:10980193

    Open questions at the time
    • Single-lab observation; nuclear/nucleolar localization has not been integrated with later plasma membrane/Golgi/endosome functions
    • Functional significance of importin-α interaction remains unclear
    • No GTP/GDP cycle characterization at this stage
  2. 2002 Medium

    Mouse knockout revealed that ARL4A contributes to spermatogenesis, establishing an in vivo physiological role distinct from simple housekeeping.

    Evidence Targeted gene disruption in mice with histological and sperm count analysis

    PMID:11909968

    Open questions at the time
    • Single-lab KO; compensatory roles of Arl4C/Arl4D not excluded
    • Molecular mechanism underlying spermatogenesis defect unknown
    • No fertility defect observed, so biological significance of reduced sperm count unclear
  3. 2007 High

    Identification of cytohesin Arf-GEFs as direct effectors established ARL4A as a plasma membrane signaling organizer that activates Arf6 independently of PI(3,4,5)P3, resolving how cytohesins reach the membrane in contexts lacking phosphoinositide signals.

    Evidence Membrane recruitment assays, direct PH-domain binding, fluorescence imaging across Arl4 family members

    PMID:17398095

    Open questions at the time
    • Relative contributions of Arl4A vs. Arl4C/Arl4D to cytohesin recruitment in specific tissues not resolved
    • Upstream signals that activate ARL4A GTP loading at the plasma membrane not identified
  4. 2011 High

    Two effector pathways were defined in the same year: GTP-dependent binding to the ELMO RBD linked ARL4A to Rac activation and actin remodeling at the plasma membrane, while GTP-dependent binding to GCC185 linked it to CLASP recruitment and Golgi maintenance, demonstrating compartment-specific effector usage.

    Evidence Two independent interaction screens, co-IP, direct binding, siRNA, Golgi fragmentation and transport assays, morphology assays with dominant-negative mutants

    PMID:21930703 PMID:22159419

    Open questions at the time
    • Whether ELMO–DOCK180 and cytohesin pathways operate simultaneously or sequentially at the plasma membrane is unknown
    • How ARL4A is partitioned between Golgi and plasma membrane pools is unresolved
    • Structural basis of GCC185 CC2 domain recognition not determined
  5. 2020 Medium

    Discovery of the ARL4A–Pak1/Pak2 interaction revealed a mutual recruitment loop at the plasma membrane that sustains Pak1 activation and cell migration, showing ARL4A is not merely upstream of Rac but also acts in parallel through Pak kinases.

    Evidence Co-IP, plasma membrane recruitment of myristoylation-deficient mutant, Arl4A depletion, fibronectin-stimulated migration assays

    PMID:31932503

    Open questions at the time
    • Single-lab study; reciprocal regulation (Pak1 recruiting Arl4A-G2A) lacks independent confirmation
    • Pak binding site on ARL4A not mapped
    • Whether Pak2 plays an equivalent functional role to Pak1 in this context is untested
  6. 2022 High

    Pak1-mediated phosphorylation at S143 was shown to recruit the chaperone HYPK, stabilizing ARL4A against proteasomal degradation and enhancing plasma membrane localization and migration — establishing post-translational protein stability as a key regulatory axis for ARL4A, distinct from classical GTPase cycling.

    Evidence Phosphoproteomics, Pak1 knockdown/overexpression, co-IP with HYPK, proteasome inhibitor rescue, S143A mutagenesis, migration assays

    PMID:35857868

    Open questions at the time
    • Whether S143 phosphorylation also influences effector binding (cytohesin, ELMO, GCC185) is unknown
    • How HYPK binding sterically or allosterically protects ARL4A from ubiquitin-proteasome targeting not determined
    • Phosphatase that reverses S143 phosphorylation not identified
  7. 2023 High

    At endosomes, ARL4A was found to bind the ESCRT-II subunit VPS36 and stabilize ESCRT-III, limiting USP8-mediated deubiquitination and thereby retarding lysosomal EGFR degradation — extending ARL4A function to receptor sorting and revealing a third subcellular site of action.

    Evidence Direct binding assays, co-IP for ESCRT-II/III, EGFR ubiquitination and degradation kinetics, USP8 recruitment assays, loss-of-function mutant

    PMID:38030597

    Open questions at the time
    • Whether ARL4A regulates sorting of receptors other than EGFR through ESCRT-II is untested
    • How ARL4A is targeted to endosomes specifically (vs. plasma membrane or Golgi) is unclear
    • GAP and GEF acting on ARL4A at the endosomal compartment are unknown

Open questions

Synthesis pass · forward-looking unresolved questions
  • The upstream GEF and GAP for ARL4A remain unidentified, leaving the regulation of its GTP loading cycle — and whether it is truly constitutively GTP-bound in vivo or subject to stimulus-dependent cycling — as a central open question.
  • No GEF or GAP identified for any Arl4 family member
  • Structural basis for effector selectivity across compartments not resolved
  • Integration of nuclear, Golgi, plasma membrane, and endosomal functions into a unified cell-biological model is lacking

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0003924 GTPase activity 4
Localization
GO:0005886 plasma membrane 4 GO:0005634 nucleus 1 GO:0005768 endosome 1 GO:0005794 Golgi apparatus 1
Pathway
R-HSA-162582 Signal Transduction 4 R-HSA-5653656 Vesicle-mediated transport 2
Partners
CYTH2DOCK1ELMO1GCC2HYPKPAK1VPS36

Evidence

Reading pass · 8 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2007 Arl4A (along with Arl4C and Arl4D) recruits cytohesin Arf-GEFs (ARNO/cytohesin-2 and relatives) to the plasma membrane by directly binding their PH domains, independently of PtdIns(3,4,5)P3 generation, thereby activating Arf6 signaling. Plasma membrane recruitment assays, binding assays between Arl4 GTPases and cytohesin PH domains, cell-based fluorescence imaging Current Biology High 17398095
2000 ARL4A localizes to nuclei and nucleoli (GDP-bound T34N mutant enriched in nucleoli), interacts with importin-alpha via its C-terminal NLS region in a nucleotide-independent manner, and its N-terminus is myristoylated. Immunofluorescence microscopy, yeast two-hybrid screening, in vitro protein-interaction assays, myristoylation assay Journal of Biological Chemistry Medium 10980193
2002 Targeted disruption of Arl4 in mice results in reduced testis weight and sperm count (~30% and ~60% reduction, respectively), indicating a role in spermatogenesis/meiosis, without affecting fertility. Targeted gene knockout in mice, histological and quantitative sperm analysis Molecular and Cellular Biology Medium 11909968
2011 Arl4A directly binds the Ras-binding domain (RBD) of ELMO proteins in a GTP-associated manner, recruiting the ELMO–DOCK180 complex to the plasma membrane, which promotes Rac activation and actin cytoskeleton remodeling (membrane ruffling, stress fiber disassembly). Two independent interaction screens (active GTPase library and brain cDNA library), biochemical binding assays, co-IP, cell-based morphology assays with dominant-negative and constitutively active mutants Journal of Biological Chemistry High 21930703
2011 Arl4A directly interacts with the Golgi golgin GCC185 in a GTP-dependent manner via the CC2 domain of GCC185, and this interaction is required for GCC185-mediated recruitment of CLASPs to the Golgi, maintenance of Golgi structure, and endosome-to-Golgi transport. Co-IP, direct binding assays (GTP-dependent), siRNA depletion, Golgi fragmentation and transport assays, CLASP recruitment assays Journal of Cell Science High 22159419
2020 Arl4A interacts with Pak1 and Pak2, recruits them to the plasma membrane, and there is mutual feedback whereby Pak1 also recruits myristoylation-deficient Arl4A-G2A to the plasma membrane; this Arl4A–Pak1 interaction (independent of Rac1 binding to Pak1) sustains Pak1 activation and promotes cell migration. Co-IP, plasma membrane recruitment assays, Arl4A-G2A mutant, Arl4A depletion, fibronectin stimulation, cell migration assays Journal of Cell Science Medium 31932503
2022 Fibronectin stimulation induces Pak1-mediated phosphorylation of Arl4A at S143 (and Arl4D at S144), which promotes binding of the chaperone HYPK to Arl4A/D, stabilizing them against proteasomal degradation and enhancing their recruitment to the plasma membrane to promote cell migration; protein stability rather than the GTPase cycle is a key regulatory mechanism for Arl4 proteins. Proteomic phosphorylation mapping, kinase identification by Pak1 knockdown/overexpression, co-IP with HYPK, proteasome inhibitor experiments, plasma membrane localization assays, cell migration assays, site-directed mutagenesis (S143A) PNAS High 35857868
2023 Endosomal Arl4A directly binds the ESCRT-II component VPS36, stabilizes VPS36–ESCRT-III association, and thereby attenuates recruitment of the deubiquitinating enzyme USP8 by CHMP2A; this prolongs EGFR ubiquitinylation and retards endosomal-to-lysosomal EGFR sorting, resulting in reduced EGFR degradation under EGF stimulation. Direct binding assays (Arl4A–VPS36 interaction), co-IP for ESCRT-II/III associations, EGFR ubiquitinylation assays, USP8 recruitment assays, EGFR degradation kinetics, loss-of-function (impaired Arl4A–VPS36 interaction mutant) Nature Communications High 38030597

Source papers

Stage 0 corpus · 8 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2007 The Arl4 family of small G proteins can recruit the cytohesin Arf6 exchange factors to the plasma membrane. Current biology : CB 105 17398095
2002 Reduced sperm count and normal fertility in male mice with targeted disruption of the ADP-ribosylation factor-like 4 (Arl4) gene. Molecular and cellular biology 50 11909968
2000 ARL4, an ARF-like protein that is developmentally regulated and localized to nuclei and nucleoli. The Journal of biological chemistry 44 10980193
2011 The Arf family GTPase Arl4A complexes with ELMO proteins to promote actin cytoskeleton remodeling and reveals a versatile Ras-binding domain in the ELMO proteins family. The Journal of biological chemistry 42 21930703
2011 ARL4A acts with GCC185 to modulate Golgi complex organization. Journal of cell science 32 22159419
2022 Phosphorylation of Arl4A/D promotes their binding by the HYPK chaperone for their stable recruitment to the plasma membrane. Proceedings of the National Academy of Sciences of the United States of America 11 35857868
2020 Cooperative recruitment of Arl4A and Pak1 to the plasma membrane contributes to sustained Pak1 activation for cell migration. Journal of cell science 11 31932503
2023 Endosomal Arl4A attenuates EGFR degradation by binding to the ESCRT-II component VPS36. Nature communications 4 38030597