Molecular interactions between ADAM12 and the non-receptor tyrosine kinase c-Src
Human ADAM12 exists in two splice variants; a long form, ADAM12-L as well as a secreted form, ADAM12-S. ADAM12-L encompasses pro-, metalloprotease, disintegrin, cysteine-rich, EGF-like, transmembrane, and cytoplasmic domains, where ADAM12-S lacks the transmembrane and cytoplasmic parts. Little is known about the structure and function of the cytoplasmic domain, but interestingly, mouse ADAM12-L has been shown to bind the proto-oncoprotein and non-receptor tyrosine kinase c-Src, raising the question whether ADAM12 and c-Src activities in cancer are linked. In order to study this in more detail, a detailed structure-function analysis of this interaction has been initiated. Using GST-pull down and competitive peptide inhibition experiments, we identified two separate binding sites in the cytoplasmic tail of human ADAM12-L that interacts with the SH3-domain of c-Src. While both sites comprise a consensus class I proline-rich Src-SH3 binding motif, a pronounced difference in binding affinity was observed, possibly due to steric availability in the 3-dimentional folding. Co-expression of ADAM12-L with wildtype c-Src and a constitutively active mutant form of c-Src lead to ADAM12-L tyrosine phosphorylation, whereas no effect of a kinase-dead c-Src mutant was detected. Surprisingly, ADAM12-mediated ecto-domain shedding of epidermal growth factor (EGF) was independent of Src-kinase activity. Current studies aim at unraveling the functional outcome of the interaction between ADAM12-L and c-Src and a potential mutual role in cancer.Acknowledgements: Grant support from the Danish Cancer Society, the Danish Medical Research Council, Novo Nordisk, Friis and Haensch Foundations.
Document Type: Abstract
Affiliations: 1: Department of Anatomy and Cell Biology, Temple University School of Medicine, Philadelphia PA, USA 2: Department of Biomedical Sciences & Biotech Research and Innovation Centre (BRIC), University of Copenhagen, Copenhagen Biocenter, Ole MaaløesVej 5, 2200 Copenhagen N, Denmark
Publication date: May 1, 2008