title: ARAP2 signals through Arf6 and Rac1 to control focal adhesion morphology.
abstract: Focal adhesions (FAs) are dynamic structures that connect the actin cytoskeleton with the extracellular matrix. At least six ADP-ribosylation factor (Arf) GTPase-activating proteins (GAPs), including ARAP2 (an Arf6 GAP), are implicated in regulation of FAs but the mechanisms for most are not well defined. Although Rac1 has been reported to function downstream of Arf6 to control membrane ruffling and cell migration, this pathway has not been directly examined as a regulator of FAs. Here we test the hypothesis that ARAP2 promotes the growth of FAs by converting Arf6·GTP to Arf6·GDP thereby preventing the activation of the Rho family GTP-binding protein Rac1. Reduced expression of ARAP2 decreased the number and size of FAs in cells and increased cellular Arf6·GTP and Rac1·GTP levels. Overexpression of ARAP2 had the opposite effects. The effects of ARAP2 on FAs and Rac1 were dependent on a functional ArfGAP domain. Constitutively active Arf6 affected FAs in the same way as did reduced ARAP2 expression and dominant negative mutants of Arf6 and Rac1 reversed the effect of reduced ARAP2 expression. However, neither dominant negative Arf6 nor Rac1 had the same effect as ARAP2 overexpression. We conclude that changes in Arf6 and Rac1 activities are necessary but not sufficient for ARAP2 to promote the growth of FAs and we speculate that ARAP2 has additional functions that are effector in nature to promote or stabilize FAs.
authors.affiliation: Laboratory of Cellular and Molecular Biology, NCI, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: Pei-Wen
authors.initials: PW
authors.lastname: Chen
authors.forename: Xiaoying
authors.initials: X
authors.lastname: Jian
authors.forename: Hye-Young
authors.initials: HY
authors.lastname: Yoon
authors.forename: Paul A
authors.initials: PA
authors.lastname: Randazzo
mesh_terms: D000089982:ADP-Ribosylation Factor 6; D020727:ADP-Ribosylation Factors / Q000378:metabolism*; D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism / Q000502:physiology*; D002448:Cell Adhesion; D002460:Cell Line; D022001:Focal Adhesions; D020690:GTPase-Activating Proteins / Q000378:metabolism / Q000502:physiology*; D005786:Gene Expression Regulation; D005982:Glutathione Transferase / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D018613:Microscopy, Confocal / Q000379:methods; D008957:Models, Genetic; D009154:Mutation; D034741:RNA, Small Interfering / Q000378:metabolism; D015398:Signal Transduction; D020830:rac1 GTP-Binding Protein / Q000378:metabolism*
chemical_list: D000089982:ADP-Ribosylation Factor 6; C446514:ARAP2 protein, human; C503237:ARFGAP1 protein, human; D002352:Carrier Proteins; D020690:GTPase-Activating Proteins; C107953:RAC1 protein, human; D034741:RNA, Small Interfering; D005982:Glutathione Transferase; D020727:ADP-Ribosylation Factors; C000715348:ARF6 protein, human; D020830:rac1 GTP-Binding Protein