3 DIFFERENT PROHORMONES YIELD A VARIETY OF HYDRA-RFAMIDE (ARG-PHE-NH2) NEUROPEPTIDES IN HYDRA-MAGNIPAPILLATA

The freshwater polyp Hydra is the most frequently used model for the s tudy of development in cnidarians, Recently we isolated four novel Arg -Phe-NH, (RFamide) neuropeptides, the Hydra-RFamides I-IV, from Hydra magnipapillata. Here we describe the molecular cloning of three differ ent preprohormones from H. magnipapillata, each of which gives rise to a variety of RFamide neuropeptides, Preprohormone A contains one copy of unprocessed Hydra-RFamide I (QWLGGRFG), II (QWFNGRFG), III/IV [(KP )HLRGRFG] and two putative neuropeptide sequences (QLMSGRFG and QLMRGR FG). Preprohormone B has the same general organization as preprohormon e A, but instead of unprocessed Hydra-RFamide III/IV it contains a sli ghtly different neuropeptide sequence [(KP)HYRGRFG]. Preprohormone C c ontains one copy of unprocessed Hydra-RFamide I and seven additional p utative neuropeptide sequences (with the common N-terminal sequence QW F/LSGRFGL). The two Hydra-RFamide II copies tin preprohormones A and B ) are preceded by Thr residues, and the single Hydra-RFamide III/IV co py tin preprohormone A) is preceded by an Asn residue, confirming that cnidarians use unconventional processing signals to generate neuropep tides from their precursor proteins. Southern blot analyses suggest th at preprohormones A and B are each coded for by a single gene, whereas one or possibly two closely related genes code for preprohormone C, N orthern blot analyses and in situ hybridizations show that the gene co ding for preprohormone A is expressed in neurons of both the head and foot regions of Hydra, whereas the genes coding for preprohormones B a nd C are specifically expressed in neurons of different regions of the head. All of this shows that neuropeptide biosynthesis in the primiti ve metazoan Hydra is already rather complex.