Characterization of nuclear localization signal sequence of CCDC84 and investigation of its interactions with histone proteins h2a1a and H2A1H

Splicing is a complex mechanism in which introns are removed from the structure of pre-mRNAs, and then mature mRNAs are formed by combining exons. More than 300 proteins can function in this mechanism. Splicing-related proteins localized in intracellular regions such as the nucleus, cytoplasm, and centrosome function in various processes such as chromatin structuring and transcription as well as regulation of splicing. In studies conducted in our department to elucidate the function of CCDC84 protein which is associated with microcephalic primordial dwarfism, it was shown that CCDC84 is localized to the nucleus and involved in the splicing mechanism, and preliminary data were obtained that it could interact with histone proteins H2A1A and H2A1H. In this thesis, it is aimed to reach data that will contribute to the further understanding of the function of CCDC84 by characterization of the nuclear localization signal (NLS) sequence of CCDC84 and testing the preliminary data for the interaction of CCDC84 with histone proteins H2A1A and H2A1H. While the characterization of the NLS sequence of CCDC84 was achieved using immunofluorescence, its potential interaction with histone proteins H2A1A and H2A1H was investigated by immunoprecipitation and immunoblot methods. As a result, a bipartite canonical NLS sequence, which is located between the 18th and 50th amino acids of CCDC84, was identified and arginine and lysine amino acids critical for the canonical nuclear transport mechanism were determined on this sequence. Possible interaction of CCDC84 with histone proteins H2A1A and H2A1H could not be confirmed and these interactions were thought to be false positives. While the findings we obtained become increasingly important with the latest studies in the literature showing the localization of CCDC84 in the centrosome, we believe that these findings will contribute significantly to the understanding of the functions of CCDC84.