Structure and promoter activity of the 5 ' flanking region of ace-1, the gene encoding acetylcholinesterase of class a in Caenorhabditis elegans

We report the structure and the functional activity of the promoter region of ace-1, the gene encoding acetylcholinesterase of class A in the nematode Caenorhabditis elegans. We found that ace-1 was trans-spliced to the SL1 s pliced leader and that transcription was initiated at a cluster of multiple starts. There was neither a TATA nor a CAAT box at consensus distances fro m these starts. Interspecies sequence comparison of the 5' regions of ace-1 in C. elegans and in the related nematode Caenorhabditis briggsae identifi ed four blocks of conserved sequences located within a sequence of 2.4 kilo bases upstream from the initiator ATG. In vitro expression of CAT reporter genes in mammalian cells allowed the determination of a minimal promoter in the first 288 nucleotides. In phenotype rescue experiments in vivo, the ac e-1 gene containing 2.4 kilobases of 5' flanking region of either C. elegan s or C. briggsae was found to restore a coordinated mobility to the uncoord inated double mutants ace-1(-); ace-2(-) of C, elegans. This showed that th e ace-1 promoter was contained in 2.4 kilobases of the 5' region, and indic ated that cis-regulatory elements as well as coding sequences of ace-1 were functionally conserved between the two nematode species. The pattern of ac e-1! expression was established through microinjection of Green Fluorescent Protein reporter gene constructs and showed a major mesodermal expression. Deletion analysis showed that two of the four blocks of conserved sequence s act as tissue-specific activators. The distal block is a mesodermal enhan cer responsible for the expression in body wall muscle cells, anal sphincte r and vulval muscle cells. Another block of conserved sequence directs expr ession in pharyngeal muscle cells pm5 and three pairs of cephalic sensory n eurons. (C) 1999 Academic Press.