Cargando…

Wee1-Regulated Apoptosis Mediated by the Crk Adaptor Protein in Xenopus Egg Extracts

Many of the biochemical reactions of apoptotic cell death, including mitochondrial cytochrome c release and caspase activation, can be reconstituted in cell-free extracts derived from Xenopus eggs. In addition, because caspase activation does not occur until the egg extract has been incubated for se...

Descripción completa

Detalles Bibliográficos
Autores principales: Smith, Jesse J., Evans, Erica K., Murakami, Monica, Moyer, Mary B., Moseley, M. Arthur, Woude, George Vande, Kornbluth, Sally
Formato: Texto
Lenguaje:English
Publicado: The Rockefeller University Press 2000
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2150666/
https://www.ncbi.nlm.nih.gov/pubmed/11134069
_version_ 1782144639100256256
author Smith, Jesse J.
Evans, Erica K.
Murakami, Monica
Moyer, Mary B.
Moseley, M. Arthur
Woude, George Vande
Kornbluth, Sally
author_facet Smith, Jesse J.
Evans, Erica K.
Murakami, Monica
Moyer, Mary B.
Moseley, M. Arthur
Woude, George Vande
Kornbluth, Sally
author_sort Smith, Jesse J.
collection PubMed
description Many of the biochemical reactions of apoptotic cell death, including mitochondrial cytochrome c release and caspase activation, can be reconstituted in cell-free extracts derived from Xenopus eggs. In addition, because caspase activation does not occur until the egg extract has been incubated for several hours on the bench, upstream signaling processes occurring before full apoptosis are rendered accessible to biochemical manipulation. We reported previously that the adaptor protein Crk is required for apoptotic signaling in egg extracts (Evans, E.K., W. Lu, S.L. Strum, B.J. Mayer, and S. Kornbluth. 1997. EMBO (Eur. Mol. Biol. Organ.) J. 16:230–241). Moreover, we demonstrated that removal of Crk Src homology (SH)2 or SH3 interactors from the extracts prevented apoptosis. We now report the finding that the relevant Crk SH2-interacting protein, important for apoptotic signaling in the extract, is the well-known cell cycle regulator, Wee1. We have demonstrated a specific interaction between tyrosine-phosphorylated Wee1 and the Crk SH2 domain and have shown that recombinant Wee1 can restore apoptosis to an extract depleted of SH2 interactors. Moreover, exogenous Wee1 accelerated apoptosis in egg extracts, and this acceleration was largely dependent on the presence of endogenous Crk protein. As other Cdk inhibitors, such as roscovitine and Myt1, did not act like Wee1 to accelerate apoptosis, we propose that Wee1–Crk complexes signal in a novel apoptotic pathway, which may be unrelated to Wee1's role as a cell cycle regulator.
format Text
id pubmed-2150666
institution National Center for Biotechnology Information
language English
publishDate 2000
publisher The Rockefeller University Press
record_format MEDLINE/PubMed
spelling pubmed-21506662008-05-01 Wee1-Regulated Apoptosis Mediated by the Crk Adaptor Protein in Xenopus Egg Extracts Smith, Jesse J. Evans, Erica K. Murakami, Monica Moyer, Mary B. Moseley, M. Arthur Woude, George Vande Kornbluth, Sally J Cell Biol Original Article Many of the biochemical reactions of apoptotic cell death, including mitochondrial cytochrome c release and caspase activation, can be reconstituted in cell-free extracts derived from Xenopus eggs. In addition, because caspase activation does not occur until the egg extract has been incubated for several hours on the bench, upstream signaling processes occurring before full apoptosis are rendered accessible to biochemical manipulation. We reported previously that the adaptor protein Crk is required for apoptotic signaling in egg extracts (Evans, E.K., W. Lu, S.L. Strum, B.J. Mayer, and S. Kornbluth. 1997. EMBO (Eur. Mol. Biol. Organ.) J. 16:230–241). Moreover, we demonstrated that removal of Crk Src homology (SH)2 or SH3 interactors from the extracts prevented apoptosis. We now report the finding that the relevant Crk SH2-interacting protein, important for apoptotic signaling in the extract, is the well-known cell cycle regulator, Wee1. We have demonstrated a specific interaction between tyrosine-phosphorylated Wee1 and the Crk SH2 domain and have shown that recombinant Wee1 can restore apoptosis to an extract depleted of SH2 interactors. Moreover, exogenous Wee1 accelerated apoptosis in egg extracts, and this acceleration was largely dependent on the presence of endogenous Crk protein. As other Cdk inhibitors, such as roscovitine and Myt1, did not act like Wee1 to accelerate apoptosis, we propose that Wee1–Crk complexes signal in a novel apoptotic pathway, which may be unrelated to Wee1's role as a cell cycle regulator. The Rockefeller University Press 2000-12-25 /pmc/articles/PMC2150666/ /pubmed/11134069 Text en © 2000 Government This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 4.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/4.0/).
spellingShingle Original Article
Smith, Jesse J.
Evans, Erica K.
Murakami, Monica
Moyer, Mary B.
Moseley, M. Arthur
Woude, George Vande
Kornbluth, Sally
Wee1-Regulated Apoptosis Mediated by the Crk Adaptor Protein in Xenopus Egg Extracts
title Wee1-Regulated Apoptosis Mediated by the Crk Adaptor Protein in Xenopus Egg Extracts
title_full Wee1-Regulated Apoptosis Mediated by the Crk Adaptor Protein in Xenopus Egg Extracts
title_fullStr Wee1-Regulated Apoptosis Mediated by the Crk Adaptor Protein in Xenopus Egg Extracts
title_full_unstemmed Wee1-Regulated Apoptosis Mediated by the Crk Adaptor Protein in Xenopus Egg Extracts
title_short Wee1-Regulated Apoptosis Mediated by the Crk Adaptor Protein in Xenopus Egg Extracts
title_sort wee1-regulated apoptosis mediated by the crk adaptor protein in xenopus egg extracts
topic Original Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2150666/
https://www.ncbi.nlm.nih.gov/pubmed/11134069
work_keys_str_mv AT smithjessej wee1regulatedapoptosismediatedbythecrkadaptorproteininxenopuseggextracts
AT evansericak wee1regulatedapoptosismediatedbythecrkadaptorproteininxenopuseggextracts
AT murakamimonica wee1regulatedapoptosismediatedbythecrkadaptorproteininxenopuseggextracts
AT moyermaryb wee1regulatedapoptosismediatedbythecrkadaptorproteininxenopuseggextracts
AT moseleymarthur wee1regulatedapoptosismediatedbythecrkadaptorproteininxenopuseggextracts
AT woudegeorgevande wee1regulatedapoptosismediatedbythecrkadaptorproteininxenopuseggextracts
AT kornbluthsally wee1regulatedapoptosismediatedbythecrkadaptorproteininxenopuseggextracts