GENES FOR MAST-CELL SERINE-PROTEASE AND THEIR MOLECULAR EVOLUTION

Trypsin-related serine proteases are encoded by a very large gene fami ly in mammals. We describe here a comparative analysis of the genomic DNA sequences of mouse, rat, and human mast-cell-specific serine prote ase genes. Strong evidence was found for multiple exchanges of genetic information between closely related members of this gene family. The 5' regulatory regions of MMCP-1 and MMCP-L share a remarkably high deg ree of sequence identity (98%), starting 10 base pairs downstream of e xon 1 and extending to the end of the presently sequenced region at po sition -1347 of the MMCP-1 gene. The remaining parts of the two genes share approximately 80% sequence identity. Evidence for at least two a dditional, but not so recent, exchanges was found in the 3' regions of the MMCP-4 and MMCP-L genes and in the 5' regions of the genes for MM CP-1 and MMCP-2. The 5' regulatory regions of all presently characteri zed mouse mast-cell-specific chymotrypsin-like serine protease genes e xhibit over 88% sequence identity in the region from the transcription initiation site to approximately position -600. An exception is MMCP- 5 which is the most distantly related member of this subfamily. The hi gh degree of sequence similarities indicates a strong evolutionary hom ogenization of the 5' regulatory region, possibly by several gene conv ersion events. In addition, several insertions of genetic information have been identified in genes for mast-cell chymases and genes for T-c ell granzymes. A number of these have been found to represent repetiti ve sequences, such as L1. The previously characterized tissue-specific enhancer element of the RMCP II gene was identified as a member of a middle repetitive sequence. A cDNA for a newly discovered pseudogene, closely related to the mouse mast cell chymases was isolated by polyme rase chain reaction amplification from a mouse connective tissue-like mast cell line. The structure of this cDNA is presented. We also prese nt the characterization of a novel spliced variant of MMCP-6 that cont ains an alternative 3' terminal exon (exon 6). The function of this va riant, if any, is still unknown. A comparative analysis of amino acid sequence identities between different hematopoietic serine proteases s hows that a high degree of sequence similarity does not always correla te with relateness in cleavage specificity. This indicates that the su bstrate specificity evolved with a higher evolutionary rate than the d egree of overall amino acid sequence identity of these proteases.