Functional interaction trap: a strategy for validating the functional consequences of tyrosine phosphorylation of specific substrates in vivo.
Protein tyrosine phosphorylation controls diverse signaling pathways, and disregulated tyrosine kinase activity plays a direct role in human diseases such as cancer. Because activated kinases exert their effects by phosphorylating multiple substrate proteins, it is difficult or impossible to assess experimentally the contribution of a particular substrate to a cellular response or activity. To overcome this problem, we have developed a novel approach termed the "functional interaction trap," in which two proteins are induced to interact in a pairwise fashion through an engineered, highly specific binding interface. We show that the functional interaction trap can be used to direct a modified tyrosine kinase to specifically phosphorylate a single substrate of choice in vivo, permitting analysis of the resulting biological output. This strategy provides a powerful tool for validating the functional significance of tyrosine phosphorylation and other post-translational modifications identified by proteomic discovery efforts.
D000818:Animals; D002460:Cell Line; D003001:Cloning, Molecular; D006801:Humans; D016350:Leucine Zippers; D051379:Mice; D015688:Oncogene Protein pp60(v-src) / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D017433:Protein Structure, Secondary; D015398:Signal Transduction; D013379:Substrate Specificity; D014443:Tyrosine / Q000378:metabolism*; D018909:src Homology Domains
D014443:Tyrosine; D015688:Oncogene Protein pp60(v-src)