DEPARTMENT OF ZOOLOGY. UNIVERSITY OF TEXAS AT AUSTIN. AUSTIN, TEXAS 78712.,
The slowpoke mutation in Drosophila produces behavioral defects and eliminates a charybdotoxin sensitive Ca2 - activated K channel (Kca). Cloning and analysis of the slowpoke (slo) gene and its homologues from other organisms demonstrated that slo encodes a K channel which is structurally and functionally homologous to the vertebrate BK-type (large conductance) K channel. Since we believe the control of BK-type channel diversity and abundance affects the signaling properties of neurons, a thorough analysis of the transcription control region should give some insight into the mechanisms regulating ion channel expression. To this end, we dissected the slowpoke promoter with nested deletions. This analysis revealed multiple promoters. More importantly, these separate promoters control the expression of alternative amino-terminal ends. Though multiple promoters control expression, two largely distinct regions control muscle and neural expression; these we now call, didactically, the muscle and neural promoters. The transcription control region is contained within an 11 kb fragment that has been shown by transgenesis to reproduce the endogenous slowpoke expression pattern. For more refined studies, we relied on evolutionary clues. We cloned and sequenced 2.2 kb and 4 kb regions of the Drosophila hydei promoter which correspond to the muscle and the neural promoters, respectively. Since D. hydei and D. melanogaster diverged over 60 million years ago, conserved elements within these control regions suggest functional relevance. Using the dual methods of deletion analysis and sequence alignment as a rapid method for analyzing a promoter, we have significantly narrowed the search for candidate cis-acting elements. In fact, the search has defined a large number of highly conserved sites interspersed with stretches of divergent sequence. Of special interest is an element within the region we have named the CNS box . Using this deletion analysis , the "CNS box" was defined as a 300 base pair region required for slo transcription in the central nervous system. Our present studies are directed towards the understanding of CNS box function.