Regulation of Apoptosis
Abstract
Apoptosis is a cellular suicide program that specifically rids an organism of damaged or superfluous cells. To define early signaling events that are required for the induction of apoptosis, we have utilized a Xenopus egg extract system capable of recapitulating several biochemical hallmarks of apoptosis in vitro. Using this in vitro egg extract system we have previously identified the adaptor protein crk as a necessary component in the apoptotic signaling pathway of these egg extracts. Furthermore, we have established that the removal of crk SH2 or N-terminal SH3 binding proteins from the extract also inhibits apoptosis. Using affinity chromatography, we have identified the major phosphotyrosine containing crk SH2 binding protein present in the egg extracts as Xenopus wee 1. Wee 1, a tyrosine kinase previously characterized as a cell cycle regulator, specifically binds to the recombinant crk SH2 domain and, furthermore, weel and crk can be co-immunoprecipitated from the egg extracts. We have found that addition of weel directly to the Xenopus egg extract accelerates apoptosis. In contrast, the addition of specific anti-weel antisera directly to the extract inhibits apoptosis. Moreover, weel exerts this pro-apoptotic function upstream of the mitochondrial requirement for apoptosis. Finally, adding recombinant Xenopus weel protein to an extract previously depleted of crk SH2 binding parmers rescues apoptosis. These data represent the first documented direct interaction between weel and an SH2 domain containing protein and is the first study to implicate a pro-apoptotic signaling capacity for the weel tyrosine kinase.
Document Details
- Document Type
- Technical Report
- Publication Date
- Sep 01, 1999
- Accession Number
- ADA381256
Entities
People
- Erica K. Evans
- Sally A. Kornbluth
Organizations
- Duke University Hospital