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PMID: 14742709 Published · ppublish English Journal Article Research Support, U.S. Gov't, P.H.S.

RNAi-mediated Hip1R silencing results in stable association between the endocytic machinery and the actin assembly machinery.

Molecular biology of the cell ·Vol. 15 ·No. 4 ·2004-04-00 ·Pages 1666-79

Engqvist-Goldstein AE, Zhang CX, Carreno S, Barroso C, Heuser JE, Drubin DG

Abstract

Actin filaments transiently associate with the endocytic machinery during clathrin-coated vesicle formation. Although several proteins that might mediate or regulate this association have been identified, in vivo demonstration of such an activity has not been achieved. Huntingtin interacting protein 1R (Hip1R) is a candidate cytoskeletal-endocytic linker or regulator because it binds to clathrin and actin. Here, Hip1R levels were lowered by RNA interference (RNAi). Surprisingly, rather than disrupting the transient association between endocytic and cytoskeletal proteins, clathrin-coated structures (CCSs) and their endocytic cargo became stably associated with dynamin, actin, the Arp2/3 complex, and its activator, cortactin. RNAi double-depletion experiments demonstrated that accumulation of the cortical actin-endocytic complexes depended on cortactin. Fluorescence recovery after photobleaching showed that dynamic actin filament assembly can occur at CCSs. Our results provide evidence that Hip1R helps to make the interaction between actin and the endocytic machinery functional and transient.

MeSH Terms
Actin Cytoskeleton/chemistry Actin-Related Protein 2 Actin-Related Protein 3 Actins/chemistry,metabolism Adaptor Proteins, Signal Transducing Blotting, Western Cell Nucleus/metabolism Cortactin Cytoskeletal Proteins/metabolism DNA/chemistry DNA-Binding Proteins/genetics Dynamins/chemistry Endocytosis Fluorescent Antibody Technique, Indirect Gene Silencing Green Fluorescent Proteins HeLa Cells Humans Luminescent Proteins/metabolism Microfilament Proteins/chemistry Microscopy, Fluorescence Phenotype Propidium/pharmacology RNA/metabolism RNA Interference RNA, Small Interfering/metabolism Time Factors Transferrin/chemistry,pharmacokinetics Vesicular Transport Proteins
Chemicals
ACTR2 protein, human ACTR3 protein, human Actin-Related Protein 2 Actin-Related Protein 3 Actins Adaptor Proteins, Signal Transducing CTTN protein, human Cortactin Cytoskeletal Proteins DNA-Binding Proteins HIP1 protein, human HIP1R protein, human Luminescent Proteins Microfilament Proteins RNA, Small Interfering Transferrin Vesicular Transport Proteins Green Fluorescent Proteins Propidium RNA DNA Dynamins
Authors & Affiliations
6 authors, click to expand affiliations / ORCID
Engqvist-Goldstein Asa E Y
Department of Molecular and Cell Biology, University of California, Berkeley, California 94720-3202, USA.
Zhang Claire X
Carreno Sebastien
Barroso Consuelo
Heuser John E
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.e03-09-0639
Published
2004-04-00
Epub
2004-00-23
Pages
1666-79
Language
English
Region
United States
NLM ID
9201390
PMCID
PMC379265
Subset
IM
Grants
NIGMS NIH HHS · R01 GM029647 · United States
NIGMS NIH HHS · R01 GM065462 · United States
NIGMS NIH HHS · GM29647 · United States
NIGMS NIH HHS · GM65462 · United States
External Links
PubMed source DOI: 10.1091/mbc.e03-09-0639 PMC Full Text View journal details
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