A MESSENGER-RNA FOR MEMBRANE FORM OF GUANYLYL CYCLASE IS EXPRESSED EXCLUSIVELY IN THE TESTIS OF THE SEA-URCHIN HEMICENTROTUS-PULCHERRIMUS

A cDNA clone encoding the membrane form of guanylyl cyclase was isolat ed from a Hemicentrotus pulcherrimus testis cDNA library and its nucle otide sequence was determined. The cDNA was 4123 bp long and an open r eading frame predicted a protein of 1125 amino acids including an appa rent signal peptide of 21 residues; a single transmembrane domain of 2 5 amino acids which divides the mature protein into an amino-terminal, extracellular domain of 485 amino acids and a carboxyl-teninal, intra cellular domain of 594 amino acids. Three potential N-linked glycosyla tion sites were present in the extracellular domain. Northern blot ana lysis of poly(A)(+)RNA from testes, ovaries, eggs and embryos at vario us developmental stages showed that the cDNA encoding guanylyl cyclase hybridized to a mRNA of 4.4 kb from the testes. We developed a large scale purification method of the phosphorylated (131 kDa) and dephosph orylated (128 kDa) forms of the membrane-bound guanylyl cyclase from H . pulcherrimus spermatozoa. The purified 131 kDa and 128 kDa forms of the guanylyl cyclase contained 26.0 +/- 1.3 and 4.3 +/- 0.7 moles of p hosphate per mot protein (mean +/- S.D.; n=6), respectively. The amino -terminal amino acids of both the 131 kDa and 128 kDa forms of the gua nylyl cyclase could not be detected, suggesting that they were blocked .