CHARACTERIZATION AND PHYLOGENETIC SIGNIFICANCE OF RHINOCEROS LUTEINIZING-HORMONE-BETA (LH-BETA) SUBUNIT MESSENGER-RNA STRUCTURE, COMPLEMENTARY-DNA SEQUENCE AND GENE COPY NUMBER

The luteinizing hormone (LH) beta subunit gene is expressed in the pit uitary glands of all mammals, whereas the closely related chorionic go nadotropin (CG) beta subunit genes have been identified only in primat es and equids, and are expressed in placenta. In the case of horses, t here is a single-copy equine (e) luteinizing hormone/chorionic gonadot ropin hormone beta subunit gene (eLH/CG beta) that (1) is expressed in both pituitary gland and placenta, (2) encodes a characteristic carbo xyl terminal peptide (CTP) extension, and (3) transcribes an atypicall y elongated 5'-untranslated region (UTR) in both pituitary and placent a. However, it is not known whether similar expression patterns and ge ne locus characteristics may be exhibited by other members of the orde r Perissodactyla (equid, rhinoceros and tapir species). To begin to in vestigate these possibilities, we undertook analysis of the rhinoceros (rn or rhino) LH/(CG?)beta gene locus and the rnLH beta cDNA. Total R NA isolated from the pituitary gland of a female white rhino was used as template for amplifying mLH beta cDNA by reverse transcription-poly merase chain reaction. Following cloning of the amplified cDNA, nucleo tide (nt) and deduced amino acid sequences were determined. The first in-frame stop codon occurred at codon position +122, suggesting that t he rnLH beta subunit does not contain a CTP. To assess gene copy numbe r, Southern blot analysis of Indian rhino genomic DNA was performed. T he resulting simple hybridization pattern indicated that, as in the ho rse and donkey, there is a single-copy gene at the mLH/(CG?)beta gene locus. Primer extension mapping of the pituitary transcriptional start site of the rnLH beta subunit gene revealed an 8 nt 5'-UTR which is s imilar to that reported for the majority of mammalian LH beta transcri pts. Northern analysis was consistent with the transcriptional start s ite findings. We postulate from these data that rhinos diverged from e quids prior to the occurrence of the mutations causing CTP expression and adoption of a non-consensus 5'-UTR/proximal promoter region. Howev er, these findings do not rule out the possibility of expression of a placental CG beta subunit lacking a CTP in rhinos. (C) 1997 Elsevier S cience B.V.