ISOLATION AND CHARACTERIZATION OF THE GENE FOR MOUSE HORMONE-SENSITIVE LIPASE

Hormone-sensitive lipase (HSL) is the rate-limiting enzyme in hydrolys is of triglycerides in adipose tissue and of cholesteryl esters in ste roidogenic tissues and macrophages. The gene encoding mouse HSL has be en isolated and characterized from two overlapping X clones, The gene spans approximately 10.4 kb and comprises 9 exons interrupted by 8 int rons. The deduced amino acid sequence specifies a protein of 759 amino acids with a Mr of 83,297 in the absence of posttranslational modific ations, The known functional domains of the HSL protein are encoded by discrete exons, with the putative catalytic site (Ser(423)) encoded b y exon 6, and the basal and regulatory phosphorylation sites (Ser(557) and Ser(559)) encoded by exon 8, In addition, a putative lipid bindin g domain occurs in exon 9. The mouse protein shows 94% identity with t he previously determined rat sequence and 85% identity with the recent ly determined human sequence. Interestingly, despite the high degree o f similarity, the three species diverge significantly for a stretch of 16 amino acid residues upstream of the phosphorylation sites, In addi tion, an error was discovered in the carboxyl-terminal portion of the previously reported rat sequence, which produced a frame shift and pre mature termination of the coding sequence, The corrected rat sequence alters the identity of 12 amino acid residues and extends the protein an additional 11 residues, We have also examined the mouse HSL gene an d 5' flanking region for nucleotide sequences that may modulate HSL ge ne transcription, Using primer extension, we identified a major transc ription initiation site 593 nucleotides upstream of the protein coding sequence. Homologies with several known gene regulatory elements are present in the HSL 5' flanking region, including sequence motifs that appear to direct expression in adipose, testis, adrenal, and myeloid t issues. (C) 1994 Academic Press, Inc.