CLONING AND EXPRESSION OF THE DOG MAST-CELL ALPHA-CHYMASE GENE

Chymases are chymotrypsin-like serine proteinases secreted by mast cel ls, alpha- and beta-chymases differ in structure, function, and mast c ell subset-and species-specific expression, Seeking genetic regulatory elements shared by alpha-chymases, we sequenced the dog alpha-gene, E xtensive homology was found in intronic and flanking sequences of the dog, human, and mouse alpha-chymase genes, but little in corresponding beta-chymase sequences, Repetitive elements probably derived from ret roposons are unique features of the dog flank, DNA blots suggest that the dog alpha-gene, like its human counterpart, may be the genome's so le chymase, unlike in rodents, in which beta-chymases predominate, Nuc lear runoff studies predict that transcriptional mechanisms explain di fferences in steady state chymase and tryptase mRNA levels between mas tocytoma and non-mast cells, In dog BR mastocytoma cells incubated wit h phorbol ester, high steady state levels of alpha-chymase mRNA drop d ramatically with little change in tryptase mRNA, whereas dexamethasone decreases expression of both mRNAs, portions of the dog or human gene 5' flank transfected into BR cells drive expression of a reporter gen e and define regions with active promoters. Thus, BR cells express hig h levels of alpha-chymase mRNA regulated independently of tryptase and support transcription using dog or human promoters, These studies rei nforce the alpha beta-chymase dichotomy and suggest the utility of BR cells in probing regulation of alpha-chymase expression.