HGNC Approved Gene Symbol: ARF6
Cytogenetic location: 14q21.3 Genomic coordinates (GRCh38): 14:49,893,082-49,897,054 (from NCBI)
ARF6 is a small GTPase that regulates membrane trafficking between endosomes and the plasma membrane (Fang et al., 2006).
Intracellular membrane trafficking involves a series of membrane budding and fusion events. These are regulated by specific cytosolic and membrane-associated protein factors, among which are a group of Ras-like small guanosine triphosphatases (GTPases) called adenosine diphosphate (ADP)-ribosylation factors (ARFs). These factors were originally identified as cofactors required for the cholera toxin-catalyzed ADP-ribosylation of Gs, alpha subunit (GNAS1; 139320); see ADP-ribosylation factor-1 (ARF1; 103180). The ARF family consists of 15 structurally related gene products that include 6 ARF proteins and 11 ARF-like proteins. The ARF proteins are divided into 3 classes on the basis of size and amino acid identity. ARF1, ARF2, and ARF3 (103190) (181 amino acids) form class I; ARF4 and ARF5 (103188) (180 amino acids) form class II; ARF6 (175 amino acids) forms class III.
Gross (2015) mapped the ARF6 gene to chromosome 14q21.3 based on an alignment of the ARF6 sequence (GenBank AF047432) with the genomic sequence (GRCh38).
D'Souza-Schorey et al. (1995) transiently expressed ARF6, ARF6 mutants, and ARF1 in Chinese hamster ovary cells and assessed the effects on receptor-mediated endocytosis. The authors demonstrated that ARF6, unlike ARF1 which is localized to the Golgi apparatus, has a central role in intra-Golgi transport, is localized to the cell periphery, and that overexpression of ARF6 causes dramatic alterations in endocytic traffic. Expression of a dominant-negative mutant of ARF6, thr27 to asn, resulted in an intracellular distribution of transferrin receptors and an inhibition of transferrin recycling to the cell surface. Cavenagh et al. (1996) examined the subcellular distribution of ARF proteins and demonstrated that ARF6 is uniquely localized to the plasma membranes in Chinese hamster ovary cells. This result suggests that ARF6 is unlikely to be involved in endocytic traffic.
Studying rat hippocampal neurons in culture, Hernandez-Deviez et al. (2002) determined that dendritic arbor development is regulated by complex interactions of ARNO (CYTH2; 602488), ARF6, and RAC1 (602048). Activation of ARNO and ARF6 resulted in signaling through RAC1 that suppressed dendritic branching.
By mass spectrometric analysis of proteins that bound a constitutively active, GTPase-deficient ARF6 mutant, Fang et al. (2006) identified the ARF GTPase-activating protein (GAP) ACAP4 (ASAP3; 616594). Immunoprecipitation analysis confirmed that ACAP4 interacted with ARF6, but not with other small GTPases examined. ACAP4 showed GAP activity with ARF6, but it did not use other small GTPases examined. For its GAP activity with ARF6, ACAP4 required membrane lipids phosphatidylinositol 4,5-bisphosphate and phosphatidic acid. Stimulation of serum-starved HeLa cells with EGF (131530) triggered redistribution of ARF6 and ACAP4 from intracellular vesicles to membrane ruffles. Membrane ruffles were extremely pronounced in cells expressing GAP-deficient ACAP4. Knockdown of ACAP4 via small interfering RNA had no effect on membrane ruffles, but it inhibited migration of HeLa cells, as determined by a wound-healing assay. Fang et al. (2006) concluded that ACAP4 functions as a GAP for ARF6 and counters the effect of ARF6 on cell membrane dynamics.
By coimmunoprecipitation studies in COS-7 cells, Falace et al. (2010) found that TBC1D24 (613577) binds ARF6 and acts as a negative regulator of ARF6. The intensity of the coimmunoprecipitated band increased in the presence of inactive GDP-locked ARF6, indicating a GDP-dependent interaction.
Zhu et al. (2012) showed that the direct, immediate, and disruptive effects of IL1-beta (IL1B; 147720) on endothelial stability in a human in vitro cell model are NF-kappa-B (see 164011)-independent and are instead the result of signaling through the small GTPase ARF6 and its activator ARNO. Moreover, Zhu et al. (2012) showed that ARNO binds directly to the adaptor protein MYD88 (602170), and thus proposed MYD88-ARNO-ARF6 as a proximal IL1-beta signaling pathway distinct from that mediated by NF-kappa-B. Finally, Zhu et al. (2012) showed that SecinH3 (182115), an inhibitor of ARF guanine nucleotide exchange factors such as ARNO, enhances vascular stability and significantly improves outcomes in animal models of inflammatory arthritis and acute inflammation.
Using N1E-115 mouse neuroblastoma cells, Torii et al. (2014) found that Ccdc120 (300947) directed neurite localization of Cyth2 and was required for Arf6 activation during differentiation. Fluorescence-tagged human CCDC120 colocalized with fluorescence-tagged Cyth2 in punctate structures within cell bodies and along extended neurite shafts and growth cones. Cyth2 inhibited ubiquitination and degradation of CCDC120, and CCDC120 targeted Cyth2 to vesicular structures. Knockdown of Ccdc120 in N1E-115 cells via small interfering RNA decreased the total number and length of neurites, dispersed Cyth2 to the cytoplasm, and reduced Arf6 activation. Torii et al. (2014) concluded that both CCDC120 and CYTH2 are required for ARF6 activation and neurite extension.
Crystal Structure
O'Neal et al. (2005) determined the 1.8-angstrom crystal structure of the cholera toxin A1 subunit in complex with human GTP-bound ARF6. Their studies revealed that the binding of the human activator elicits dramatic changes in CTA1 loop regions that allow nicotinamide adenine dinucleotide (NAD+) to bind to the active site. The extensive toxin:ARF-GTP interface surface mimics the ARF-GTP recognition of normal cellular protein partners, which suggests that the toxin has evolved to exploit promiscuous binding properties of ARFs.
Cavenagh, M. M., Whitney, J. A., Carroll, K., Zhang, C., Boman, A. L., Rosenwald., A. G., Mellman, I., Kahn, R. A. Intracellular distribution of Arf proteins in mammalian cells. J. Biol. Chem. 271: 21767-21774, 1996. [PubMed: 8702973] [Full Text: https://doi.org/10.1074/jbc.271.36.21767]
D'Souza-Schorey, C., Li, G., Colombo, M. I., Stahl, P. D. A regulatory role for ARF6 in receptor-mediated endocytosis. Science 267: 1175-1178, 1995. [PubMed: 7855600] [Full Text: https://doi.org/10.1126/science.7855600]
Falace, A., Filipello, F., La Padula, V., Vanni, N., Madia, F., De Pietri Tonelli, D., de Falco, F. A., Striano, P., Dagna Bricarelli, F., Minetti, C., Benfenati, F., Fassio, A., Zara, F. TBC1D24, an ARF6-interacting protein, is mutated in familial infantile myoclonic epilepsy. Am. J. Hum. Genet. 87: 365-370, 2010. [PubMed: 20727515] [Full Text: https://doi.org/10.1016/j.ajhg.2010.07.020]
Fang, Z., Miao, Y., Ding, X., Deng, H., Liu, S., Wang, F., Zhou, R., Watson, C., Fu, C., Hu, Q., Lillard, J. W., Jr., Powell, M., Chen, Y., Forte, J. G., Yao, X. Proteomic identification and functional characterization of a novel ARF6 GTPase-activating protein, ACAP4. Molec. Cell. Proteomics 5: 1437-1449, 2006. Note: Erratum: Molec. Cell. Proteomics 5: 1718 only, 2006. [PubMed: 16737952] [Full Text: https://doi.org/10.1074/mcp.M600050-MCP200]
Gross, M. B. Personal Communication. Baltimore, Md. 3/31/2015.
Hernandez-Deviez, D. J., Casanova, J. E., Wilson, J. M. Regulation of dendritic development by the ARF exchange factor ARNO. Nature Neurosci. 5: 623-624, 2002. [PubMed: 12032543] [Full Text: https://doi.org/10.1038/nn865]
O'Neal, C. J., Jobling, M. G., Holmes, R. K., Hol, W. G. J. Structural basis for the activation of cholera toxin by human ARF6-GTP. Science 309: 1093-1096, 2005. [PubMed: 16099990] [Full Text: https://doi.org/10.1126/science.1113398]
Torii, T., Miyamoto, Y., Tago, K., Sango, K., Nakamura, K., Sanbe, A., Tanoue, A., Yamauchi, J. Arf6 guanine nucleotide exchange factor cytohesin-2 binds to CCDC120 and is transported along neurites to mediate neurite growth. J. Biol. Chem. 289: 33887-33903, 2014. [PubMed: 25326380] [Full Text: https://doi.org/10.1074/jbc.M114.575787]
Zhu, W., London, N. R., Gibson, C. C., Davis, C. T., Tong, Z., Sorenson, L. K., Shi, D. S., Guo, J., Smith, M. C. P., Grossmann, A. H., Thomas, K. R., Li, D. Y. Interleukin receptor activates a MYD88-ARNO-ARF6 cascade to disrupt vascular stability. Nature 492: 252-255, 2012. [PubMed: 23143332] [Full Text: https://doi.org/10.1038/nature11603]