Transcriptional cofactors play complex roles in developmental gene regulation. The evolutionarily conserved transcriptional corepressor C-terminal binding protein (CtBP) is recruited by a variety of transcriptional factors that play crucial roles in development and disease; this protein contains a central NAD(H)-binding core domain homologous to D2 hydroxy acid dehydrogenase enzymes, as well as an unstructured C-terminal domain. NAD(H) binding is important for CtBP repression activity and enables CtBP to function as a metabolic sensor to regulate gene expression through cellular metabolic status. The striking homology of CtBP to metabolic dehydrogenase enzymes and the presence of an enzymatic activity in CtBP also raise the possibility of a direct link between cellular metabolism and transcriptional regulation. The unstructured C-terminus is susceptible to modifications by sumoylation and phosphorylation, which can influence CtBP subcellular localization and stability. In this study, we established genetic rescue assays to determine how the enzymatic activity and the C-terminal domain of CtBP function in the context of Drosophila melanogaster development. The mutant phenotypes and specific gene regulatory effects indicate that both the catalytic site of CtBP and the C-terminal domain play important roles in development. Our results indicate that the structural and enzymatic features of CtBP, previously thought to be dispensable for overall transcriptional control, are critical for modulating this protein's activity in diverse developmental settings.In addition, we identified hundreds of potential CtBP target genes affected by the enzymatic activity and the C-terminal domain of CtBP by genome-wide transcriptome analysis. We show that these enzymatic and structural features regulate multiple cellular processes such as metamorphosis and metabolism. Particularly, we show that CtBP may regulate Drosophila innate immunity, a potential novel function implied by CtBP. Taken together, our results may open doors to discover novel functions of CtBP in multiple biological processes.
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