부산대학교 도서관

cabut (cbt) encodes a transcription factor involved in Drosophila dorsal closure (DC), and it is expressed in embryonic epithelial sheets and yolk cells during this process upon activation of the Jun N-terminal kinase (JNK) signaling pathway. Additional studies suggest that cbt may have a role in multiple developmental processes, such as neuroendocrine cell remodeling, ecdysone response, circadian rhythm, axon guidance, synaptogenesis, cell growth and cell cycle. The vertebrate orthologes of Cbt, the family of TIEG transcription factors, are involved in proliferation and cancer. In this thesis, we have generated different genetic, biochemical and immunological tools for the study of the function of Cbt during fly development, its transcriptinal targets and the structural domains responsible for its function. To analyze Cbt subcellular localization throughout embryogenesis, as well as Cbt´s transcriptional targets, we generated a Cbt-specific antibody that allowed us to (1) detect new Cbt expression patterns and (2) perform Chromatin Immunoprecipitation assays followed by microarray analyses (ChIP-on-chip). Immunohistochemical analyses during DC revealed that Cbt is expressed in the yolk, epidermal and amnioserosal cells, which contribute to the DC process. Also, Cbt is expressed in the central and peripheral nervous system. Moreover, cell culture assays have determinated that the 71PNKKPRL77 motif of Cbt functions as a nuclear localization signal (NLS), and that the conserved TIEG´s NLS, is not functional in insects. Direct mutagensis demonstrated that K73 and K74 residues are necessary for Cbt transport to the nucleus. Whole embryo microarray analyses of cbt mutants during DC were performed, and they revealed that Cbt regulates more than 1000 genes during the DC process, such as the 20-hydroxyecdysone hormone (20E)-inducible genes ImpE1 and ImpL1. Indeed, 20E regulates Cbt in the epidermis. Finally, ChIP-on-chip studies showed direct binding of Cbt to genomic regions close to Cbt and NPC1a (Niemann-Pick Type C-1) genes, which regulate trafficking of sterols through endocytic/secretory pathway. Taken together, these results suggest that 20E and lipid trafficking are involved in the DC through Cbt.