Neither gene locus nor gene sequence characterizations have been reported for the β subunits of guinea pig (gp) LH and putative gp chorionic gonadotropin (CG). Descriptions of this locus would allow comparison with functionally relevant molecular genetic features of other species' homologous loci including the single-copy equid LH/CGβ gene and the primate LHβ-CGβ gene cluster locus. Contiguous cDNA and genomic DNA fragments spanning the entire mature coding sequence of gpLHβ mRNA, gpCGβ mRNA and a homologous gpLH/CGβ gene were amplified using PCR methodologies. With the exception of one silent mutation, the two cDNA and the genomic sequences were identical where they overlapped. Comparison of guinea pig coding sequence with LHβ, CGβ and LH/CGβ sequences of other vertebrate species revealed the following order of similarity expressed as percent coding sequence identity: rhinoceros LHβ (83·6%)>pig LHβ (81·8%)>donkey LH/CGβ=bovine LHβ (81·5%)> horse LH/CGβ (80·6%)>dog LHβ (79·7%)>human LHβ (78·2%)>rat LHβ (77·9%)>human CGβ (75·8%)>turkey LHβ (52·7%); values that are generally consistent with recently postulated phylogenetic relationships. Like the consensus mammalian LHβ gene, the 5′-flanking region of the gpLH/CGβ gene contains a single TATA sequence 37 bp upstream of the translation start codon. The first in-frame stop codon occurred at codon position + 122 which is consistent with the 121 amino acid residue length of the consensus mammalian mature LHβ peptide. To estimate gene copy number, full-length gpLHβ cDNA was radiolabeled and hybridized to Southern blots of guinea pig genomic DNA digested with a panel of six restriction endonucleases. The resulting simple hybridization pattern strongly suggested that there is a single-copy gpLH/CGβ gene. Northern analysis of total pituitary RNA using the same probe indicated that gpLHβ transcript size is indistinguishable from that of consensus mammalian pituitary LHβ mRNAs (Ο750 nucleotides). Despite amplifying gpCGβ from placental RNA, positive signal was not detected in Northern blot lanes containing guinea pig total RNA prepared from placentae collected at three gestational ages (17·3 days, 24·3 days and 68 days (term)). Other data suggest that inability to detect Northern blot signal could have been due to low relative tissue concentrations of gpCGβ transcript and/or sampling at gestational time-points that missed peak periods of mRNA expression. We conclude that, with respect to gene copy number, coding sequence and pituitary mRNA size, the gpLH/CGβ gene locus reflects the CTP-less consensus mammalian LHβ condition. However, based on the capacity of this single-copy gene to express in both pituitary and placental tissues, gpLH/CGβ also exhibits functional similarities with the single-copy equine. LH/CGβ locus.
ASJC Scopus subject areas
- Molecular Biology