Distinct variant DNA-binding sites determine cell-specific autoregulated expression of the Drosophila POU domain transcription factor drifter in midline glia or trachea

Transcriptional regulators utilizing the POU domain DNA-binding motif have been shown to form multi-protein complexes dependent on the POU domain itself and its flexible recognition of various octamer sequence elements. We have identified two variant POU domain recognition elements DFRE1 and DFRE2, which are found within a 514-bp autoregulatory enhancer of the Drosophila melanogaster POU domain gene drifter (dfr). Both elements are capable of binding bacterially produced full-length DFR protein with high affinity, although they differ in the 5'-to-3' orientation of POU-specific and POU homeodomain subelements. When placed in dfr loss-of-function genetic backgrounds, all expression of dfr-lacZ fusion genes under control of the autoregulatory enhancer is dependent on DFR activity levels. However, the complete enhancer sequence directs beta-galactosidase expression in only a subset of cells which normally express the endogenous DFR protein, including the middle pair of midline glias of the ventral nerve cord, the oenocyte clusters, and all tracheal cells. In addition, DFRE1 and DFRE2 exhibit separable tissue-specific functions when independently disrupted or deleted. Disruption of DFRE1 function specifically abolishes beta-galactosidase expression in the middle pair of midline glias. Deletion of DFRE causes a specific loss of tracheal expression, leaving oenocyte and midline glia expression intact. These results suggest that dfr cell-specific autoregulation is determined by the context of DFR POU domain binding within the enhancer, which is possibly mediated by the formation of recognition element-specific heteromultimeric complexes containing additional tissue-specific factors.