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KMID : 0620920100420030216
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Experimental & Molecular Medicine
2010 Volume.42 No. 3 p.216 ~ p.222
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A double point mutation in PCL-γ1 (Y509A/F510A) enhances Y783 phosphorylation and inositol phospholipid-hydrolyzing activity upon EGF stimulation
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Chung Sang-Hee
Kim Sung-Kuk
Kim Jung-Kuk
Yang Yong-Ryoul
Suh Pann-Ghill
Chang Jong-Soo
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Abstract
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Growth factor stimulation induces Y783 phosphorylation of phosphoinositide-specific PLC-γ1, and the subsequent activation of this enzyme in a cellular signaling cascade. Previously, we showed that a double point mutation, Y509A/F510A, of PLC-γ1, abolished interactions with translational elongation factor 1-α. Here, we report that the Y509A/F510A mutant PLC-γ1 displayed extremely high levels of Y783 phosphorylation and enhanced catalytic activity, compared to wild-type PLC-γ1, upon treatment of COS7 cells with EGF. In quiescent COS7 cells, the Y509A/F510A mutant PLC-γ1 exhibited a constitutive hydrolytic activity, whereas the wild-type counterpart displayed a basal level of activity. Upon treatment of COS7 cells with EGF, the Y783F mutation in Y509A/F510A PLC-γ1 (Y509A/F510A/Y783F triple mutant) cells also led to an enhanced catalytic activity, whereas Y783F mutation alone displayed a basal level of activity. Our results collectively suggest that the Y509A/F510A mutant is more susceptible to receptor tyrosine kinase-induced Y783 phosphorylation than is wild-type PLC-γ1, but no longer requires Y783 phosphorylation step for the Y509A/F510A mutant PLC-γ1 activation in vivo.
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KEYWORD
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phosphatidylinositol 4, 5-bisphosphate, phospholipase Cγ, protein-tyrosine kinases, src homology domains
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