PISA Technology : a Novel Tool to Select Intrabodies to Target the Post-Translationally Modified Proteome in the Cell
The complexity of the proteome greatly outnumbers that of the genome and the diversity is mainly due to post-translational modifications (PTMs). Indeed, PTMs are fundamental regulators of many pathways and processes. Modifications like phosphorylation, acetylation, ubiquitination or methylation regulate several intracellular processes such as cell cycle progression, chromatin structure and gene expression or signal transduction cascades by activating or deactivating enzymes. PTMs also determine different protein conformations, which have a great physiological and pathological relevance. Currently no general strategy allows to interfere directly with a post-translationally modified protein, versus its unmodified counterpart. Thus, no FDA approved drugs is directed against a protein PTM. Antibodies represent a desirable tool for targeting PTMs proteins with high specificity. Intrabodies (intracellular antibodies) are small antibody variable domains that target intracellular proteins and are stable in the cell cytosol. An overview on the preparation of recombinant antibodies libraries and on the Intracellular Antibody Capture Technology (IACT) will be presented along with, a new method that we recently developed, the PISA (Post-translational Intracellular Silencing Antibody) technology. This technology allows to systematically generate intrabodies that are able to target and neutralize specific PTMs of proteins in mammalian cells. In particular, the application of PISA to selectively interfere with site-specific acetylated residues of histones will be described, a new selective strategy for epigenetic modulation.