Studies of the interaction of novel forms of the p185(c-neu) receptor ectodomain

We are exploring the roles of the extracellular domain (ECD) of the rat p185c-neu, one member of the erbB family transmembrane receptor tyrosine kinases, which features a signal diversifying mechanism. This mechanism involves the formation of various homo- and hetero-oligomers. One set of the rat c-neu extracellular subdomain individually-deleted constructs and their corresponding NR6 transfectants with or without the coexpression of other erbB family members were studied for inter-receptor interactions. A c-neu-IgG(human) immunoadhesin was also developed to search for the rat p185c-neu ECD specific interacting proteins and to generate a rabbit polyclonal antibody, Anti-NEX. We found that the rat c-neu extracellular subdomain deletions significantly limited cell surface targeting of these mutant receptors, especially in single transfectants. The deletions at the c-neu different extracellular subdomains resulted in distinct changes of the mutant c-neu receptors\u27 phosphotyrosine (PY) content. Extracellular treatment with Anti-NEX significantly down-modulated p185c-neu and increased the PY content of the remaining p185 c-neu. Rat E18 frozen sections stained by high concentrations of the c-neu-IgG(human) immunoadhesin showed a pattern parallel to p185c-neu tissue distribution pattern. Two lines of the c-neu-IgG(human) transgenic driven by the human cytomegalovirus promoter/enhancer were established. RT-PCR assay detected the transcripts of the c-neu-IgG(human) transgene and the mouse c-neu in all examined organs. The c-neu-IgG(human) protein was also detectable in the plasma. But the c-neu-IgG(human) transgenic mice lacked any obvious phenotype. These studies reveal novel regulatory roles of the p185 c-neu ECD. The results of this dissertation may provide new thoughts for cancer therapy and protein engineering