2 HIGHLY HOMOLOGOUS RIBONUCLEASE GENES EXPRESSED IN MOUSE EOSINOPHILSIDENTIFY A LARGER SUBGROUP OF THE MAMMALIAN RIBONUCLEASE SUPERFAMILY

Two putative ribonucleases have been isolated from the secondary granu les of mouse eosinophils, Degenerate oligonucleotide primers inferred from peptide sequence data were used in reverse transcriptase-PCR reac tions of bone marrow-derived cDNA. The resulting PCR product was used to screen a C57BL/6J bone marrow cDNA library, and comparisons of repr esentative clones showed that these genes and encoded proteins are hig hly homologous (96% identity at the nucleotide level; 92/94% identical /similar at the amino acid level), The mouse proteins are only weakly homologous (approximate to 50% amino acid identity) with the human eos inophil-associated ribonucleases (i.e., eosinophil-derived neurotoxin and eosinophil cationic protein) and show no sequence bias toward eith er human protein, Phylogenetic analyses established that the human and mouse loci shared an ancestral gene, but that independent duplication events have occurred since the divergence of primates and rodents. Th e duplication event generating the mouse genes was estimated to have o ccurred <5 x 10(6) years ago (versus 30 to 40 x 10(6) years ago in pri mates), The identification of independent duplication events in two ex tant mammalian orders suggests a selective advantage to having multipl e eosinophil granule ribonucleases. Southern blot analyses in the mous e demonstrated the existence of three additional highly homologous gen es (i.e., five genes total) as well as several more divergent family m embers, The potential significance of this observation is the implicat ion of a larger gene subfamily in primates (i.e., humans).