dCtBP-dependent and -independent repression activities of the Drosophila knirps protein

Transcriptional repressor proteins play essential roles in controlling the correct temporal and spatial patterns of gene expression in Drosophila mela nogaster embryogenesis. Repressors such as Knirps, Kruppel, and Snail media te short-range repression and interact with the dCtBP corepressor. The mech anism by which short-range repressors block transcription is not well under stood; therefore, we have undertaken a detailed structure-function analysis of the Knirps protein. To provide a physiological setting for measurement of repression, the activities of endogenous or chimeric Knirps repressor pr oteins were assayed on integrated reporter genes in transgenic embryos. Two distinct repression functions were identified in Knirps. One repression ac tivity depends on dCtBP binding, and this function maps to a C-terminal reg ion of Knirps that contains a dCtBP binding motif. In addition, an N-termin al region was identified that represses in a CtBP mutant background and doe s not bind to the dCtBP protein in vitro. Although the dCtBP protein is imp ortant for Knirps activity on some genes, one endogenous target of the Knir ps protein, the even-skipped stripe 3 enhancer, is not derepressed in a CtB P mutant. These results indicate that Knirps can utilize two different path ways to mediate transcriptional repression and suggest that the phenomenon of short-range repression may be a combination of independent activities.