THE CLONING OF A CAENORHABDITIS-ELEGANS GUANYLYL CYCLASE AND THE CONSTRUCTION OF A LIGAND-SENSITIVE MAMMALIAN NEMATODE CHIMERIC RECEPTOR/

Substantial guanylyl cyclase activity was detected in membrane fractio ns prepared from Caenorhabditis elegans (100 pmol cGMP/min/mg at 20 de grees C or 500 pmol cGMP/min/mg art 37 degrees C), suggesting the pote ntial existence of orphan cyclase receptors in the nematode, Using deg enerate primers, a cDNA clone encoding a putative membrane form of the enzyme (GCY-X-1) was obtained, The apparent cyclase was most closely related to the mammalian natriuretic peptide receptor family, and reta ined cysteine residues conserved within the extracellular domain of th e mammalian receptors, Expression of the cDNA in COS-7 cells resulted in low, but detectable guanylyl cyclase activity (about 2-fold above v ector alone), The extracellular and protein kinase homology domain of the mammalian receptor (GC-B) for C-type natriuretic peptide (CNP) was fused to the catalytic domain of GCY-X, and expressed in COS-7 cells to determine whether ligand dependent regulation would now be obtained . The resulting chimeric protein (GC-BX1) was active, and CNP elevated cGMP in a concentration-dependent manner, Subsequently, a search of t he genome data base demonstrated the existence of at least 29 differen t genes From C. elegans that align closely with the catalytic domain o f GCY-X-1, and thus an equally large number of different regulatory li gands may exist.