Visit our Expo - Redox and Inflammation signaling 2012

Session X : Cell death in cancer Poster X, 98
Inhibition of cell-free apoptosis in Xenopus oocyte extracts by Src kinase
Alexander A. Tokmakov1, Mikako Shirouzu1, Shigeyuki Yokoyama1,2,3
1Genomic Sciences Center, RIKEN Yokohama Institute, Tsurumi, Yokohama, 230-0045
Japan, 2RIKEN Harima Institute at Spring-8, 3University of Tokyo, Japan. E-mail:
tokmak@gsc.riken.go.jp
Oncoproteins can induce both cell proliferation and apoptosis, depending on cellular context.
v-Src oncoprotein has been shown to activate several kinases, which can inhibit apoptosis,
including ERK and PI3-K. To further characterize the involvement of Src kinase in apoptosis,
we investigated the effect of Src overexpresion in the cell-free extracts prepared from oocytes
and eggs of the African clawed frog, Xenopus laevis. These extracts can recapitulate a range
of apoptotic events, including release of cytochrom C, caspase activation, and nuclear
fragmentation (Newmeyer et al., 1994). Notably, interphase-arrested and metaphase-arrested
extracts have different susceptibility to undergo apoptosis. The MAPK pathway active in
metaphase-arrested extracts was shown to protect them from apoptosis (Tashker et al., 2002).
In the present study we measured the caspase 3/7 activation and cytochrom C release in
apoptotic cell-free Xenopus oocyte and egg extracts, overexpressing constitutively active (v-
Src) and kinase negative (Src KN) mutants of Src kinase. We found that interphase-arrested
oocyte extracts were highly apoptotic. They released cytochrom C and displayed elevated
levels of caspase activity after several hours’-long incubation at room temperature. Caspase
activation in these extracts could be prevented by overexpression of either v-Src or Src KN
protein kinase mutants. No or little effect of Src overexpression on cytochrom C release was
detected. On the other hand, metaphase arrested egg extracts were resistant to apoptosis and
they could not be regulated by Src kinase. Our data suggest that the inhibition of apoptosis in
Xenopus oocyte extracts by Src kinase is not mediated by activation of the MAPK pathway.
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