Mecanisms of regulation of the expression of the atypical protein kinase C PKMz and identification of interacting proteins in a neuro specific context.

Abstract

Breast cancer is a frequent disease in women, and presents a heterogeneous clinical behavior, being composed by an array of molecular profiles. In this context, the protein kinase C family, a group of ten serine/threonine kinases, is of extreme importance, being related with several cellular functions. However, the exact role of each isoform is not fully understood in all the subtypes of this disease. The identification of the differential activity, subcellular localization and substrate characterization of each specific isoform in specific breast cancer subtypes is, therefore, essential for a better understanding of breast cancer biology.Our group recently developed monoclonal and polyclonal antibodies which specifically recognize active classical PKCs (cPKCs), and with this innovative tool in hands we propose to study the cPKCs activity in a breast cancer cell line, MCF7, representative of luminal A positive estrogen receptor subtype. We are also going to utilize this tool in samples from luminal A patients. Previous data from our group showed that PKCsbI and bII are the major nuclear isoforms in MCF7 cells. We are now going to detect the possible nuclear substrates of these enzymes through Bimolecular Fluorescence Complementation method, and also through immunoprecipitation. Still, we are going to observe the phosphorylation in specific sites of histones, which could act as important epigenetic marks, by performing in vitro assays and chromatin immunoprecipitation, validating the data in samples from luminal A patients. This way, we are going to i