We are interested in understanding how myogenic lineages are specified during early embryonic development. Since
hlh-1 is the earliest known marker of striated muscle lineages, we are using
hlh-1 control elements to investigate striated muscle specification. Previous deletion analysis of the
hlh-1 promoter region suggested the presence of multiple lineage specific control elements [Krause et al., 1994]; at the time these studies were done, current techniques for further characterization of these control elements (gfp and the minimal
pes-10 promoter) were not available. As the first step towards identifying the critical components in specifying striated muscle lineages, we have begun to characterize in more detail the
hlh-1 control elements that mediate early expression in myogenic precusors of the C, D, and MS lineages. Fragments covering the
hlh-1 upstream region were tested for the ability to activate a minimal promoter (the deleted
pes-10 upstream region). We have broken the upstream region into a number of functional segments, and have so far focused on two active segments of 170 and 120nt which are adjacent and approximately 2.3kb upstream of the gene. Either segment can direct reporter expression in muscle precursors of the D and C lineages. The latter segment also shows enhancer activity in the MS granddaughter cells. We are in the process of defining minimal sequences of these enhancers for each of the observed expression patterns. Once the smallest enhancer elements are defined, we plan to take two approaches to identify factors that control the activities of these elements. We will carry out genetic screens in parallel to identify factors that are functioning in the pathway for lineage specific specification of striated muscle. Toward the same goal, we will also use the identified
hlh-1 control elements as biochemical probes to search for interacting factors.