Molecular Characterization of Human MUC16 (CA125) in Breast Cancer.

摘要

This study was designed to understand the role and implications of MUC16 cytoplasmic tail in breast cancer pathogenesis. We would like to update our findings with respect to it since our last report submission. One important question towards this end was whether MUC16 indeed undergoes cleavage, which was addressed using a dual-epitope tagging (N-ter FLAG and C-ter HA) (last report). Having established this, we wanted to identify the potential cleavage site on MUC16, for which we generated a series of deletion constructs suggesting this cleavage might be in the proximal juxta-membrane region. In addition, we have demonstrated that MUC16 undergoes ubiquitylation in the cytoplasmic lysine (K) residue and N-glycosylation in the extracellular aspargine (N) that are responsible its stability. In an effort to identify any intracellular trigger for the cleavage of MUC16 (e.g. phosphorylation on the tyrosine (Y), serine (S) and/or threonine (T)), we have carried out SDM of these residues, but none of them was able to prevent the cleavage of MUC16. While further biochemical characterizations are being carried out, to gauge at the functional significance we have generated stable MCF7 cells and the functional studies are being carried out. The significance of this study is manifold: (i) This will enable us to establish the precise site of cleavage, cellular location of cleavage and the trigger (we do not know at this point whether it is induced or constitutive), which is very much necessary in order to design any kind of therapeutics, (ii) post-translational modifications affecting the trafficking and stability and interaction(s) of it are of immense importance to understand the functional mechanism(s).