Expression and genomic organization of a medaka fish novel membrane form of guanylyl cyclase/orphan receptor

A novel membrane guanylyl cyclase (membrane GC), OIGC8, was identified in the medaka fish Oryzias latipes by the isolation of full-length cDNA (4958 bp) and genomic DNA (14.3 kbp) clones. Phylogenetic analysis indicated that OIGC8 does not belong in any known vertebrate membrane GC sub-family. OIGC8 consists of an extracellular domain (214 residues), a transmembrane segment (19 residues), and an intracellular protein kinase-like domain (284 residues) and a cyclase catalytic domain (228 residues), although the extracellular domain is about half the length (around 450 residues) of other known vertebrate membrane GCs. OIGC8 transiently expressed in COS-7 cells exhibited only basal guanylyl cyclase activity. None of the known ligands (rat ANP, BNP, CNP, and C-ANF) and various medaka fish tissue extracts, which activated OIGC1, OIGC2, and OIGC7 differentially, stimulated basal activity, suggesting that OIGC8 is an orphan receptor. The OIGC8 gene consists of 24 exons and exists as a single copy on the medaka fish genome. Northern blot hybridization showed that a 5 kb-OIGC8 mRNA was expressed in the kidney and the testis at a high level and a 3.3 kb-OIGC8 mRNA was expressed only in the brain. The RNase protection, RNA Ligase-Mediated Rapid Amplification of cDNA Ends (RLM-RACE), and reverse transcription-polymerase chain reaction (RT-PCR) analyses demonstrated that the 3.3 kb-OIGC8 mRNA detected in the brain is transcribed from the second transcription initiation site, and contains an intron at the position prior to the catalytic domain, the translation product of which appears to be a protein lacking the cyclase catalytic domain.