Identification of signalling proteins interacting with B-Raf in the yeast two-hybrid system

被引:0
|
作者
Papin, C [1 ]
Denouel, A [1 ]
Calothy, G [1 ]
Eychene, A [1 ]
机构
[1] CTR UNIV ORSAY,INST CURIE,UMR 146 CNRS,LAB 110,F-91405 ORSAY,FRANCE
关键词
B-Raf; c-Raf-1; Ras; MEK-1; MEK-2; 14-3-3; MAP kinase cascade;
D O I
暂无
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Recent studies suggested the existence of Ras/B-Raf/MEK-1 complexes and a critical role for B-Raf in regulating the MAP kinase/ERKs signalling pathway, We report, here, that both Ras and MEK-1 proteins interact physically with B-Raf proteins in the yeast two-hybrid system, In addition, by screening a mouse brain cDNA library, we isolated additional B-Raf interacting proteins, These include three members of the 14-3-3 proteins family (eta, theta and zeta) and the MEK-2 protein, We also show that c-Raf-1, previously reported to interact with beta and zeta 14-3-3 proteins, also interacts with eta and theta 14-3-3 proteins in the two-hybrid system, By using different portions of the B-Raf protein, we mapped the regions of the protein involved in these interactions, Specifically, we have characterized B-Raf specific sequences required for an efficient interaction with MER proteins, We show that, consequently, B-Raf interacts with MEK-1 and MEK-2 with a better affinity than does c-Raf-1, thus strengthening the notion that B-Raf is a stronger MER activator than c-Raf-1, Our results also suggest that a MEK specific sequence, not present in MAP kinase kinases which are not activated by members of the Raf family, is required for the interaction with Raf proteins.
引用
收藏
页码:2213 / 2221
页数:9
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