EFA6 facilitates the assembly of the tight junction by coordinating an Arf6-dependent and -independent pathway.
We have previously reported that EFA6, exchange factor for Arf6, is implicated upon E-cadherin engagement in the process of epithelial cell polarization. We had found that EFA6 acts through stabilization of the apical actin ring onto which the tight junction is anchored. Mutagenesis experiments showed that both the catalytic domain of EFA6 and its C-terminal domain were required for full EFA6 function. Here we address the contribution of the specific substrate of EFA6, the small G protein Arf6. Unexpectedly, depletion of Arf6 by RNA interference or expression of the constitutively active fast-cycling mutant (Arf6T157N) revealed that Arf6 plays an opposing role to EFA6 by destabilizing the apical actin cytoskeleton and the associated tight junction. However, in complementation experiments, when the C-terminal domain of EFA6 is co-expressed with Arf6T157N, it reverts the effects of Arf6T157N expressed alone to faithfully mimic the phenotypes induced by EFA6. In addition, we find that the two signaling pathways downstream of EFA6, i.e. the one originating from the activated Arf6GTP and the other one from the EFA6 C-terminal domain, need to be tightly balanced to promote the proper reorganization of the actin cytoskeleton. Altogether, our results indicate that to regulate the tight junction, EFA6 activates Arf6 through its Sec7 catalytic domain as it modulates this activity through its C-terminal domain.
Klein
Stéphanie
S
Institut de Pharmacologie Moléculaire et Cellulaire, CNRS-UMR 6097, Université de Nice Sophia-Antipolis, 660 route des Lucioles, 06560 Valbonne, France.
Partisani
Mariagrazia
M
Franco
Michel
M
Luton
Frédéric
F
D000089982:ADP-Ribosylation Factor 6; D020727:ADP-Ribosylation Factors / Q000378:metabolism*; D000818:Animals; D020134:Catalytic Domain; D006224:Cricetinae; D003599:Cytoskeleton / Q000378:metabolism; D004285:Dogs; D005786:Gene Expression Regulation; D051379:Mice; D008954:Models, Biological; D009419:Nerve Tissue Proteins / Q000378:metabolism / Q000502:physiology*; D010641:Phenotype; D017434:Protein Structure, Tertiary; D034622:RNA Interference; D015398:Signal Transduction; D019108:Tight Junctions / Q000378:metabolism*
D000089982:ADP-Ribosylation Factor 6; D009419:Nerve Tissue Proteins; D020727:ADP-Ribosylation Factors; C000715727:Arf6 protein, mouse
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