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PMID: 17151356 Published · ppublish English Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't

Negative regulation of yeast Eps15-like Arp2/3 complex activator, Pan1p, by the Hip1R-related protein, Sla2p, during endocytosis.

Molecular biology of the cell ·Vol. 18 ·No. 2 ·2007-02-00 ·Pages 658-68

Toshima J, Toshima JY, Duncan MC, Cope MJ, Sun Y, Martin AC, Anderson S, Yates JR, Mizuno K, Drubin DG

Abstract

Control of actin assembly nucleated by the Arp2/3 complex plays a crucial role during budding yeast endocytosis. The yeast Eps15-related Arp2/3 complex activator, Pan1p, is essential for endocytic internalization and proper actin organization. Pan1p activity is negatively regulated by Prk1 kinase phosphorylation after endocytic internalization. Phosphorylated Pan1p is probably then dephosphorylated in the cytosol. Pan1p is recruited to endocytic sites approximately 25 s before initiation of actin polymerization, suggesting that its Arp2/3 complex activation activity is kept inactive during early stages of endocytosis by a yet-to-be-identified mechanism. However, how Pan1p is maintained in an inactive state is not clear. Using tandem affinity purification-tagged Pan1p, we identified End3p as a stoichiometric component of the Pan1p complex, and Sla2p, a yeast Hip1R-related protein, as a novel binding partner of Pan1p. Interestingly, Sla2p specifically inhibited Pan1p Arp2/3 complex activation activity in vitro. The coiled-coil region of Sla2p was important for Pan1p inhibition, and a pan1 partial loss-of-function mutant suppressed the temperature sensitivity, endocytic phenotypes, and actin phenotypes observed in sla2DeltaCC mutant cells that lack the coiled-coil region. Overall, our results establish that Sla2p's regulation of Pan1p plays an important role in controlling Pan1p-stimulated actin polymerization during endocytosis.

MeSH Terms
Actin-Related Protein 2-3 Complex/metabolism Actins/metabolism Adaptor Proteins, Vesicular Transport/metabolism Carrier Proteins/genetics,metabolism DNA-Binding Proteins/metabolism Endocytosis/genetics Fungal Proteins/analysis,antagonists & inhibitors,metabolism Gene Deletion Microfilament Proteins Mutation Saccharomyces cerevisiae Proteins Saccharomycetales/chemistry,physiology,ultrastructure
Chemicals
Actin-Related Protein 2-3 Complex Actins Adaptor Proteins, Vesicular Transport Carrier Proteins DNA-Binding Proteins Fungal Proteins Microfilament Proteins PAN1 protein, S cerevisiae Saccharomyces cerevisiae Proteins
Authors & Affiliations
10 authors, click to expand affiliations / ORCID
Toshima Jiro
Department of Molecular and Cell Biology, University of California, Berkeley, CA 94720-3202, USA.
Toshima Junko Y
Duncan Mara C
Cope M Jamie T V
Sun Yidi
Martin Adam C
Anderson Scott
Yates John R
Mizuno Kensaku
Drubin David G
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Article Info
Journal
Molecular biology of the cell
Abbr.
Mol Biol Cell
ISSN
1059-1524
DOI
10.1091/mbc.e06-09-0788
Published
2007-02-00
Epub
2006-00-06
Pages
658-68
Language
English
Region
United States
NLM ID
9201390
PMCID
PMC1783767
Subset
IM
Grants
NIGMS NIH HHS · R01 GM042759 · United States
NIGMS NIH HHS · R37 GM042759 · United States
NIGMS NIH HHS · GM50399 · United States
NIGMS NIH HHS · GM42759 · United States
NIGMS NIH HHS · R01 GM050399 · United States
External Links
PubMed source DOI: 10.1091/mbc.e06-09-0788 PMC Full Text View journal details
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