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The F-BAR protein Syp1 negatively regulates WASp-Arp2/3 complex activity during endocytic patch formation.


ABSTRACT: Actin polymerization by Arp2/3 complex must be tightly regulated to promote clathrin-mediated endocytosis. Although many Arp2/3 complex activators have been identified, mechanisms for its negative regulation have remained more elusive. To address this, we analyzed the yeast arp2-7 allele, which is biochemically unique in causing unregulated actin assembly in vitro in the absence of Arp2/3 activators.We examined endocytosis in arp2-7 mutants by live-cell imaging of Sla1-GFP, a coat marker, and Abp1-RFP, which marks the later actin phase of endocytosis. Sla1-GFP and Abp1-RFP lifetimes were accelerated in arp2-7 mutants, which is opposite to actin nucleation-impaired arp2 alleles or deletions of Arp2/3 activators. We performed a screen for multicopy suppressors of arp2-7 and identified SYP1, an FCHO1 homolog, which contains F-BAR and AP-2micro homology domains. Overexpression of SYP1 in arp2-7 cells slowed Sla1-GFP lifetimes closer to wild-type cells. Further, purified Syp1 directly inhibited Las17/WASp stimulation of Arp2/3 complex-mediated actin assembly in vitro. This activity was mapped to a fragment of Syp1 located between its F-BAR and AP-2micro homology domains and depends on sequences in Las17/WASp outside of the VCA domain.Together, these data identify Syp1 as a novel negative regulator of WASp-Arp2/3 complex that helps choreograph the precise timing of actin assembly during endocytosis.

SUBMITTER: Boettner DR 

PROVIDER: S-EPMC2828323 | biostudies-literature | 2009 Dec

REPOSITORIES: biostudies-literature

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The F-BAR protein Syp1 negatively regulates WASp-Arp2/3 complex activity during endocytic patch formation.

Boettner Douglas R DR   D'Agostino Jessica L JL   Torres Onaidy Teresa OT   Daugherty-Clarke Karen K   Uygur Aysu A   Reider Amanda A   Wendland Beverly B   Lemmon Sandra K SK   Goode Bruce L BL  

Current biology : CB 20091203 23


<h4>Background</h4>Actin polymerization by Arp2/3 complex must be tightly regulated to promote clathrin-mediated endocytosis. Although many Arp2/3 complex activators have been identified, mechanisms for its negative regulation have remained more elusive. To address this, we analyzed the yeast arp2-7 allele, which is biochemically unique in causing unregulated actin assembly in vitro in the absence of Arp2/3 activators.<h4>Results</h4>We examined endocytosis in arp2-7 mutants by live-cell imaging  ...[more]

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