MOLECULAR-CLONING AND CHARACTERIZATION OF A NOVEL TISSUE-SPECIFIC CALPAIN PREDOMINANTLY EXPRESSED IN THE DIGESTIVE-TRACT

In the course of the genomic cloning of nCL-2, a stomach-specific calp ain, we identified a genomic clone encoding a novel member of the calp ain large subunit family and designated it 'nCL-4'. First, using exon sequences, we cloned the cDNA for mouse nCL-4. Based on this sequence, we also cloned the cDNAs for rat and human nCL-4. In the case of huma n nCL-4, the longest open reading frame encodes 690 amino acid residue s (M-r 79095) with equal sequence similarities (50-55%) to both ubiqui tous and organ-specific calpain large subunits from mammals. The deduc ed amino acid sequence revealed that nCL-4 is highly conserved among m ammals. nCL-4 can be aligned without significant deletions or insertio ns, and, thus, like other calpains, can be divided into four domains ( I-IV). The significant similarity of domains II and IV to those in con ventional calpain large subunits suggests the potential protease activ ity and Ca2+-binding ability of nCL-4. Northern blot analysis revealed that the mRNA for nCL-4 is expressed predominantly in stomach and sma ll intestine but not in uterus, suggesting specialized functions of nC L-4 in the digestive tract. When overexpressed in COS-7 cells, a speci fic band for nCL-4 was detected. In addition, the gene coding for nCL- 4 was localized on human chromosome 1.