Document Detail

Live-cell imaging RNAi screen identifies PP2A-B55alpha and importin-beta1 as key mitotic exit regulators in human cells.
MedLine Citation:
PMID:  20711181     Owner:  NLM     Status:  MEDLINE    
When vertebrate cells exit mitosis various cellular structures are re-organized to build functional interphase cells. This depends on Cdk1 (cyclin dependent kinase 1) inactivation and subsequent dephosphorylation of its substrates. Members of the protein phosphatase 1 and 2A (PP1 and PP2A) families can dephosphorylate Cdk1 substrates in biochemical extracts during mitotic exit, but how this relates to postmitotic reassembly of interphase structures in intact cells is not known. Here, we use a live-cell imaging assay and RNAi knockdown to screen a genome-wide library of protein phosphatases for mitotic exit functions in human cells. We identify a trimeric PP2A-B55alpha complex as a key factor in mitotic spindle breakdown and postmitotic reassembly of the nuclear envelope, Golgi apparatus and decondensed chromatin. Using a chemically induced mitotic exit assay, we find that PP2A-B55alpha functions downstream of Cdk1 inactivation. PP2A-B55alpha isolated from mitotic cells had reduced phosphatase activity towards the Cdk1 substrate, histone H1, and was hyper-phosphorylated on all subunits. Mitotic PP2A complexes co-purified with the nuclear transport factor importin-beta1, and RNAi depletion of importin-beta1 delayed mitotic exit synergistically with PP2A-B55alpha. This demonstrates that PP2A-B55alpha and importin-beta1 cooperate in the regulation of postmitotic assembly mechanisms in human cells.
Michael H A Schmitz; Michael Held; Veerle Janssens; James R A Hutchins; Otto Hudecz; Elitsa Ivanova; Jozef Goris; Laura Trinkle-Mulcahy; Angus I Lamond; Ina Poser; Anthony A Hyman; Karl Mechtler; Jan-Michael Peters; Daniel W Gerlich
Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't     Date:  2010-08-15
Journal Detail:
Title:  Nature cell biology     Volume:  12     ISSN:  1476-4679     ISO Abbreviation:  Nat. Cell Biol.     Publication Date:  2010 Sep 
Date Detail:
Created Date:  2010-09-02     Completed Date:  2010-10-22     Revised Date:  2013-12-19    
Medline Journal Info:
Nlm Unique ID:  100890575     Medline TA:  Nat Cell Biol     Country:  England    
Other Details:
Languages:  eng     Pagination:  886-93     Citation Subset:  IM    
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MeSH Terms
Cell Nucleus Division / drug effects,  physiology
Chromosomes / metabolism
Cyclin-Dependent Kinases / antagonists & inhibitors,  metabolism
Flavonoids / pharmacology
Golgi Apparatus / metabolism
HeLa Cells
Histones / metabolism
Image Processing, Computer-Assisted / methods
Interphase / physiology
Leupeptins / pharmacology
Microscopy, Confocal / methods
Microscopy, Fluorescence / methods
Mitosis / drug effects,  physiology*
Models, Biological
Phosphorylation / physiology
Piperidines / pharmacology
Protein Binding / physiology
Protein Phosphatase 2 / genetics,  metabolism*
RNA Interference*
RNA, Small Interfering / genetics
Spindle Apparatus / metabolism
beta Karyopherins / genetics,  metabolism*
Grant Support
037538//Wellcome Trust; 073980//Wellcome Trust; 081361//Wellcome Trust; 083524//Wellcome Trust; 097945//Wellcome Trust; C08577//Biotechnology and Biological Sciences Research Council; C12944//Biotechnology and Biological Sciences Research Council; G0301131//Medical Research Council; G0801738//Medical Research Council
Reg. No./Substance:
0/Flavonoids; 0/Histones; 0/KPNB1 protein, human; 0/Leupeptins; 0/PPP2R1A protein, human; 0/PPP2R2A protein, human; 0/Piperidines; 0/RNA, Small Interfering; 0/beta Karyopherins; 133407-82-6/benzyloxycarbonylleucyl-leucyl-leucine aldehyde; 45AD6X575G/alvocidib; EC Kinases; EC protein, human; EC Phosphatase 2

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