Molecular cloning and RNA expression of a novel Drosophila calpain, Calpain C.
The calpains are Ca(2+)-activated cysteine proteases whose biochemical properties have been extensively characterized in vitro. Less is known, however, about the physiological role of calpains. In this respect, Drosophila melanogaster is a useful experimental organism to study calpain activity and regulation in vivo. The sequencing of the fly genome has been recently completed and a novel calpain homologue has been identified in the CG3692 gene product. We embarked on the cloning and characterization of this putative novel calpain. We demonstrate that the actual calpain is different from the predicted protein and we provide experimental evidence for the correction of the genomic annotation. This novel protein, Calpain C, must be catalytically inactive, having mutated active site residues but is otherwise structurally similar to the other known fly calpains. Moreover, we analysed Calpain C RNA expression during Drosophila development by RT-PCR and RNA in situ hybridization, which revealed strong expression in the salivary glands.
Spadoni
Cesare
C
Institute of Enzymology, Biological Research Center, Hungarian Academy of Sciences, P.O. Box 7, Budapest H-1518, Hungary.
Farkas
Attila
A
Sinka
Rita
R
Tompa
Peter
P
Friedrich
Peter
P
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002154:Calpain / Q000096:biosynthesis* / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D004247:DNA / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D029721:Drosophila Proteins / Q000096:biosynthesis* / Q000235:genetics*; D004331:Drosophila melanogaster / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D017403:In Situ Hybridization; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009841:Oligonucleotides / Q000737:chemistry; D012313:RNA / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D013997:Time Factors
D018076:DNA, Complementary; D029721:Drosophila Proteins; D009841:Oligonucleotides; D012333:RNA, Messenger; D012313:RNA; D004247:DNA; D002154:Calpain; C473990:calpain C, Drosophila; D002118:Calcium