Protocadherinlerin memeli hücrelerinde moleküler analizi

Cadherins are a big family of transmembrane, cell-cell adhesion proteins and protocadherins constitute the largest subgroup of cadherins. Although cadherins are extensively studied, less information is available about protocadherins. Protocadherins have been shown to function in cell adhesion and they are thought to be the molecular markers of neuronal identity. Also, recent studies revealed the involvement of protocadherins in cancer and neurodegenerative diseases. We aimed to characterize a protocadherin, which showed a cell cycle dependent subcellular localization. We generated various mutant versions of the protocadherin and analyzed subcellular localizations of wild type and mutated proteins throughout the cell cycle using microscopy. We also investigated the role of the cytoplasmic domain in regulation of cell cycle dependent localization of the protein. In order to identify the interaction partners of the cytoplasmic domain, we used two orthogonal approaches: 1. GST-fusion pull-down assay followed by mass spectrometry; 2. BioID method (proximity-dependent biotin identification combined with biotin-streptavidin affinity chromatography and mass spectrometry). These two experimental strategies helped us to discover interaction partner candidates of the protocadherin. Furthermore, we performed bead aggregation assay. We demonstrated that the protocadherin that we studied does not have homophilic adhesion activity. This research project produced novel knowledge about the protocadherins and showed the promise of protocadherins to be used in cancer diagnosis and treatment.