Nucleotide polymorphisms, transcriptional analysis, gene expression of the bovine growth hormone secretagogue receptor 1a (GHS-R1a) gene and its genetic association with growth and carcass traits in cattle

Ghrelin - Growth hormone secretagogue receptor 1a (GHS-R1a) is involved in many important functions including growth hormone (GH) secretion and food intake. In this chapter, we explore existing nucleotide polymorphisms, transcriptional analysis, gene expression of the bovine GHS-R1a gene and its genetic association with growth and carcass traits in cattle. Firstly, we evaluated haplotype variety and characterized the microsatellite ((TG)n, 5'-untransrated region (UTR)) and nucleotide polymorphisms of the GHS-R1a gene in cattle. Nucleotide sequencing of this gene (~6 kb) revealed 47 single nucleotide polymorphisms (SNPs), four indels and the microsatellite ((GTTT)n, Intron 1). The 19 haplotypes were constructed from all nucleotide viability patterns and divided into 3 major groups. Four SNPs (L24V, nt456(G>A), D191N and nt667(C>T)) and DelR242 in Exon 1 and a haplotype block of about 2.2 kb (nt667(C>T) ~ nt2884 (A>G)) were identified in Bos taurus breeds. Significant breed differences in allele frequencies of the two microsatellites, nt-7(C>A), L24V, and DelR242 loci were found. A DelR242 was found in the Japanese Shorthorn (frequency: ~ 0.44), Japanese Brown, 5 European cattle breeds, the Philippine native cattle, but none detected in the Japanese Black nor the Mishima Island cattle. Secondly, 5'-rapid amplification of cDNA ends (5'-RACE) and reverse transcription-polymerase chain reaction (RT-PCR) analyses revealed that there were two different kinds of transcripts: spliced, without a microsatellite within 5'-UTR (GHS-R1a); and non-spliced, with the microsatellite (GHS-R1b). Thirdly, we examined age-related changes in the expressions of GHS-R1a and GHS-R1b (the truncated-type receptor) in the arcuate nucleus, pituitary gland and other tissues by real-time RT-PCR in cattle. The GHS-R 1a mRNA expression in the arcuate nucleus of post-weaning calves was more than 10-fold higher than those of pre-weaning calves and cows, and its expression level was the highest in all tissues examined. The GHS-R1a mRNA expression in the pituitary gland of pre-weaning calves was higher than those of post-weaning calves and cows. The GHS-R1b mRNA expression was widespread in all tissues examined and predominantly occurred in the pancreas, pituitary gland, spleen and arcuate nucleus in adult. Fourthly, we carried out a genetic association study between five nucleotide polymorphisms (5'UTR microsatellite ((TG)n), nt-7(C>A), L24V, DelR242 and Intron 1 microsatellite (GTTT)n) of the GHS-R1a gene and growth and carcass traits in 1,285 steers sired by 117 Japanese Black bulls in a progeny testing program. Statistical analysis revealed that the 5'UTR microsatellite locus had a significant additive effect on carcass weight (CW) and average daily gain (ADG). One of the four major microsatellite alleles (19-TG allele) with an allele frequency of 0.145, had a significantly desirable effect on CW and ADG. We proposed a translational hypothesis that the association is due to differences in the secondary structure of GHS-R1b mRNA (the non-spliced type with the 5'UTR microsatellite) among the GH-SR1a gene haplotypes. Finally, we predicted the potential increase in profitability due to increased CW in cow-calf fattening enterprises through planned matings based on DNA testing of the 5'UTR microsatellite. We concluded that the 19-TG allele could potentially be an economically useful nucleotide marker for growth and carcass traits in Japanese Black cattle.