Deciphering the origin of Met-enkephalin and Leu-enkephalin in Lobe-finnedfish: cloning of Australian lungfish proenkephalin

The previous detection of Met-enkephalin and Leu-enkephalin in the CNS of t he Australian lungfish, Neoceratodus forsteri, in a molar ratio comparable to mammals suggested that the lungfish proenkephalin precursor should conta in the sequences of both Met-enkephalin and Leu-enkephalin as seen for mamm alian proenkephalin. However, the cloning of a full-length proenkephalin cD NA from the CNS of the Australian lungfish indicates that the organization of this precursor is more similar to amphibian proenkephalin than mammalian proenkephalin. The Australian lungfish cDNA is 1284 nucleotides in length and the open reading frame (267 amino acids) contains seven opioid sequence s (GenBank #AF232671). There are five copies of the Met-enkephalin sequence flanked by sets of paired basic amino acid proteolytic cleavage sites and two C-terminally extended forms of Met-enkephalin: YGGFMRSL and YGGFMGY. As seen for amphibians, no Leu-enkephalin sequence was detected in the Austra lian lungfish proenkephalin cDNA. The fact that Leu-enkephalin has been ide ntified by radioimmunoassay and HPLC analysis in the CNS of the Australian lungfish indicates that a Leu-enkephalin-coding gene, distinct from proenke phalin, must be expressed in lungfish. Potential candidates may include a p rodynorphin- or other opioid-like gene. Furthermore. the absence of a Leu-e nkephalin sequence in lungfish and amphibian proenkephalin would suggest th at the mutations that yielded this opioid sequence in tetrapod proenkephali n occurred at some point in the radiation of the amniote vertebrates. (C) 2 000 Elsevier Science B.V. All rights reserved.