CLONING AND BIOCHEMICAL-CHARACTERIZATION OF THE CYCLOPHILIN HOMOLOGS FROM THE FREE-LIVING NEMATODE CAENORHABDITIS-ELEGANS

Cyclosporin A (CsA) is the most widely used immunosuppressive agent, w hose properties are exerted via an interaction with cyclophilin, resul ting in down-regulation of signal-transduction events in the T-cell. C yclophilin is identical with peptidylprolyl cis-trans isomerase (PPI; EC 5.2.1.8), an enzyme which catalyses the isomerization between the t wo proline conformations in proteins, thereby acting as a catalyst in protein-folding events. Several reports indicate that CsA has potent a nti-parasitic activity, effective against both protozoan and helminth species. In order to understand the various biological roles that cycl ophilins play we have initiated a study of these proteins in the genet ically tractable nematode Caenorhabditis elegans. Here we describe the cloning and characterization of 11 cyclophilin genes (cyp-1 to -11) d erived from this nematode; this is currently the greatest number of is oforms described in a single species. Southern blotting and physical m apping indicated that these genes are dispersed throughout the nematod e genome. A high degree of conservation exists between several isoform s, which also share characteristics with the ubiquitous isoforms previ ously described. The remaining isoforms are divergent, having altered CsA-binding domains and additional non-cyclophilin domains, which may impart compartmental specificity. Ten of these isoforms have been expr essed in Escherichia coli, and the resultant fusion proteins have been examined biochemically for PPI activity, which they all possess, Isom erase activity is highest in the conserved and lowest in divergent iso forms, perhaps indicating a more specific substrate for the latter. An alysis of the C. elegans cyp genes will provide answers as to the role s played by cyclophilins in protein folding and signal transduction.