CLONING OF A CDNA-ENCODING A PUTATIVE MOLT-INHIBITING HORMONE FROM THE EYESTALK OF THE SAND SHRIMP METAPENAEUS-ENSIS

Degenerate primers were designed from the amino acid sequence of the n europeptide Pej-SGP-IV of the shrimp Penaeus japonicus. Reverse transc riptase-polymerase chain reaction (RT-PCR) was performed using eyestal k complementary DNA of the sand shrimp Metapenaeus ensis. A partial cD NA that codes for a protein homologous to the neuropeptide Pej-SG-IV w as cloned. The partial cDNA was used as a probe to screen the eyestalk cDNA library. Several cDNA clones with nucleotide sequence identical to the partial cDNA were isolated. The largest cDNA is 957 bp with an open reading frame consisting of a coding sequence 315 bp in length. T he deduced amino acid of the neuropeptide consists of 77 amino acids a nd is preceded by a signal peptide of 28 amino acids. Because the dedu ced amino acid sequence of the shrimp cDNA is highly homologous to the Pej-SGP-IV of P. japonicus (which is molt inhibiting) and to other cr ustaceans' molt-inhibiting hormones (MIHs), the shrimp neuropeptide is tentatively called MeMIH. Northern blot analysis and RT-PCR showed th at MeMIH is expressed in the postmolt, intermolt, and premolt stages o f the shrimp eyestalks and the brain. Moreover, RNA message can also b e detected in the nervous tissues of newly developed larvae. MeMIH is, however, not found in the muscle, swimming leg, and hepatopancreas. R esults from genomic Southern blot analysis and amplification of the sh rimp genomic DNA by polymerase chain reaction (PCR) suggest that a sin gle copy of the MIH gene is present in the genome. The structural orga nization of the gene for the shrimp putative MIH is similar to that of the crab Charybdis feriatus.