Growth Hormone Locus Research Articles

The valuable heterosis and genetic inheritance of live weight trait by growth hormone gene action in Ongole-grade cattle

The crucial genetic factors of live weight inheritance including breeding value, dominance deviation, heritability and heterosis had not been studied in Ongole-grade cows. This study aims to identify the genotypic components including breeding value, dominance deviation, heritability and heterosis of live weight trait in Ongole-grade cows. A total of 78 blood samples from parental cows and 2 blood samples from parental Ongole-breed bulls were collected to be analyzed for presences of growth hormone (GH) genes. Presences of GH locus in blood samples were screened using PCR-RFLP method involving restricted enzyme Msp1 on agarose-gel (1.2%). Data were analyzed by statistical program in Excel XP. Results showed that the population mean (μ) of cow live weight was 385.98± 2.49 kg. Genetic component inheritance of homozygous genotypes of GH +/+ and GH –/– and heterozygous genotype of GH +/– were dominated by dominance gene action, rather than the additive gene action resulting the valuable heterosis for live weight trait of 3.2 percent. The heritability of dominance gene action (hD 2 ) was 0.96 categorized as higher heritability, while heritability of additive gene action (hA 2 ) was 0.04 categorized as low heritability under the equilibrium frequencies of both GH + (p = 0.48) and GH – (q = 0.52) genes without considering the nongenetic causes of environmental effect in the population of cow live weight.

Gene Content and Coding Diversity of the Growth Hormone Loci of Apes.

The growth hormone (GH) locus has experienced a dramatic evolution in primates, becoming multigenic and diverse in anthropoids. Despite sequence information from a vast number of primate species, it has remained unclear how the multigene family was favored. We compared the structure and composition of apes' GH loci as a prerequisite to understanding their origin and possible evolutionary role. These thorough analyses of the GH loci of the chimpanzee, gorilla, and orangutan were done by resorting to previously sequenced bacterial artificial chromosomes (BACs) harboring them, as well as to their respective genome projects data available in GenBank. The GH loci of modern man, Neanderthal, gibbon, and wild boar were retrieved from GenBank. Coding regions, regulatory elements, and repetitive sequences were identified and compared among species. The GH loci of all the analyzed species are flanked by the genes CD79B (5') and ICAM-1 (3'). In man, Neanderthal, and chimpanzee, the loci were integrated by five almost indistinguishable genes; however, in the former two, they rendered three different hormones, and in the latter, four different proteins were derived. Gorilla exhibited six genes, gibbon seven, and orangutan four. The sequences of the proximal promoters, enhancers, P-elements, and a locus control region (LCR) were highly conserved. The locus evolution might have implicated duplications of the ancestral pituitary gene (GH-N) and subsequent diversification of the copies, leading to the placental single GH-V gene and the multiple CSH genes.

Associations between polymorphisms in the growth hormone locus and reproductive efficiency in dairy cattle in Pujon

Selection using genetic markers (MAS) is carried out in an effort to accelerate livestock breeding programs, through molecular studies individual superior livestock can be detected earlier so that the selection program can be directed and controlled. This study aims to analyze genetic diversity at the growth hormone locus that is thought to have a relationship with reproductive and production traits in Friesian Holstein Crossed (FHC) dairy cattle through the polymerase chain reaction - restriction fragment length polymorphisms (PCR-RFLP) method. The study was conducted in Pujon subdistrict using 53 FHC cattle samples. The results showed that at the GH gene locus only one genotype was found, AA with an allele frequency of 1.00. Based on these results, it was concluded that the GH gene in FHC cattle is monomorphic. The information in this study can be used as a reference in formulating livestock breeding programs.

Growth hormone locus polymorphism in Ongole Grade and Fries Holland Grade bulls and its implications in the digestibility of feed

This research aimed to analyze polymorphism of the growth hormone (GH) gene and its implication on digestibility of low and high quality feed. Forty bulls Ongole Grade (OG) and 30 bulls Fries Holland Grade (FHG) had been selected and subjected for an experiment to measure body weight gain, dry matter intake, feed efficiency and feed digestibility. The treatment was crude fibrous feed from various plants with a fibre weight of 15% (P1: low quality) and 22.5% (P2: high quality). The 389 bp of GH gene was amplified (F = 5'-CCCACGGGAGAGAATGAGGC–3'; dan R = 5'-TGAGGAACTGCAGGGGGPA-3') and digested using Msp1 restriction enzyme. Polymorphism between OC and HFC bulls was 45%. The FHG bulls' weight gain were significantly higher than of OC (P<0.05) at first to third week, but become similar (P>0.05) at fourth week. It was concluded that the polymorphism between FHG and OC was high and HFC have a growth response and feed efficiency higher than those of OC.

SAT-303 3D Mapping of the Human Growth Hormone Locus Identifies Putative Regulatory Hubs for Genes Involved in Cellular Signalling and Cancer-Related Pathways

Growth hormone (GH) is a peptide hormone predominantly produced in the pituitary that is crucial for normal growth and metabolism. Downstream effects of GH are mediated through binding to the GH receptor (GHR) and consequent activation of key signalling cascades including JAK-STAT, MAPK, PI3K-Akt and mTOR. The GH locus is comprised of five evolutionarily related genes under the control of an upstream locus control region which coordinates tissue-specific expression of these genes in the pituitary and placenta (1). Compromised GH signalling and genetic variation in these genes has been implicated in various disorders including cancer. We hypothesised that polymorphisms which occur within the GH locus have the potential to impact on disease phenotypes by altering or disrupting gene regulation. We used the CoDeS3D (Contextualize Developmental SNPs using 3D Information) algorithm to analyse 529 common single nucleotide polymorphisms (SNPs) across the locus. This algorithm identifies colocalised Hi-C and eQTL associations to determine which SNPs are associated with a change in gene expression at loci that physically interact within the nucleus. We identified 181 common SNPs that interacted with 292 eGenes in 48 different tissues. 145 eGenes were regulated in trans. We performed pathway enrichment of identified eGenes and found these to be enriched in GH/GHR-related downstream cellular signalling pathways including MAPK, PI3K-Akt-mTOR, and ErbB signalling. Enrichment was also observed in the Wnt and Hippo signalling pathways. There was also a significant representation of these eGenes in pathways associated with hepatocellular, colorectal, breast and non-small cell lung carcinoma. 33 eQTL SNPs identified in our study were found to be of regulatory importance in a genome-wide Survey of Regulatory Elements (SuRE) reporter screen (2). In addition, 7 eQTL SNPs were located in known enhancer regions. Our data suggests that regions within the GH locus form regulatory hubs for multiple genes in cis and trans (intra and inter-chromosomal), many of which are involved in mediating GH function in normal and pathogenic states.Reference: (1) Tsai et al. Nucleic Acids Res 2016, 44, 10, 4651 (2) van Arensbergen et al. Nat Genet 2019, 51, 7, 1160.

3D interactions with the growth hormone locus in cellular signalling and cancer-related pathways.

Growth hormone (GH) is a peptide hormone predominantly produced by the anterior pituitary and is essential for normal growth and metabolism. The GH locus contains five evolutionarily related genes under the control of an upstream locus control region that coordinates tissue-specific expression of these genes. Compromised GH signalling and genetic variation in these genes has been implicated in various disorders including cancer. We hypothesised that regulatory regions within the GH locus coordinate expression of a gene network that extends the impact of the GH locus control region. We used the CoDeS3D algorithm to analyse 529 common single nucleotide polymorphisms (SNPs) across the GH locus. This algorithm identifies colocalised Hi-C and eQTL associations to determine which SNPs are associated with a change in gene expression at loci that physically interact within the nucleus. One hundred and eighty-one common SNPs were identified that interacted with 292 eGenes across 48 different tissues. One hundred and forty-five eGenes were regulated in trans. eGenes were found to be enriched in GH/GHR-related cellular signalling pathways including MAPK, PI3K-AKT-mTOR, ERBB and insulin signalling, suggesting that these pathways may be co-regulated with GH signalling. Enrichment was also observed in the Wnt and Hippo signalling pathways and in pathways associated with hepatocellular, colorectal, breast and non-small cell lung carcinoma. Thirty-three eQTL SNPs identified in our study were found to be of regulatory importance in a genome-wide Survey of Regulatory Elements reporter screen. Our data suggest that the GH locus functions as a complex regulatory region that coordinates expression of numerous genes in cis and trans, many of which may be involved in modulating GH function in normal and disease states.

Evolution of IFN subgroups in bony fish - 2. analysis of subgroup appearance and expansion in teleost fish with a focus on salmonids

A relatively large repertoire of type I interferon (IFN) genes is apparent in rainbow trout/Atlantic salmon, that includes six different IFN subgroups (IFNa-IFNf) belonging to the three known type I IFN groups (1–3) in bony fish. Whether this is true for other salmonids, and how the various type I subgroups evolved in teleost fish was studied using the extensive genomic resources available for fish. This confirmed that salmonids, at least the Salmoninae, indeed have a complex (in terms of IFN subgroups present) and large (number of genes) IFN repertoire relative to other teleost fish. This is in part a consequence of the salmonid 4 R WGD that duplicated the growth hormone (GH) locus in which type I IFNs are generally located. Divergence of the IFN genes at the two GH loci was apparent but was not seen in common carp, a species that also underwent an independent 4 R WGD. However, expansion of IFN gene number can be found at the CD79b locus of some perciform fish (both freshwater and marine), with expansion of the IFNd gene repertoire. Curiously the primordial gene order of GH-IFNc-IFNb-IFNa-IFNe is largely retained in many teleost lineages and likely reflects the tandem duplications that are taking place to increase IFN gene number. With respect to the evolution of the IFN subgroups, a complex acquisition and/or loss has occurred in different teleost lineages, with complete loss of IFN genes at the GH or CD79b locus in some species, and reduction to a single IFN subgroup in others. It becomes clear that there are many variations to be discovered regarding the mechanisms by which fish elicit protective (antiviral) immune responses.

Polymorphisms of the CAPN, CAST, LEP, GH, GHR, IGF-1 and MSTN loci of Colombian Creole hair x Pelibuey sheep sheep crossbreeds

The objective of this study was to characterize single nucleotide polymorphisms at the Calpain (CAPN), Calpastatin (CAST), Leptin (LEP), Growth Hormone (GH), Growth Hormone Receptor (GHR), Insulin-Like Growth Factor 1 (IGF-1) and myostatin (MSTN) loci in Columbian Creole hair sheep crossed with Pelibuey sheep. In 192 individuals, the CAST and MSTN loci were genotyped by PCR-RFLP, and the CAPN, GH, GHR, IGF-1 and LEP loci by PCR-SSCP and sequences. The following parameters were calculated: allelic and genotypic frequencies, observed (Ho) and expected (He) heterozygosity, fixation index and deviations from Hardy-Weinberg equilibrium (HWE). In the CAPN locus, higher frequencies of the T allele (70%) were found with respect to C (30%) and the frequencies of the TT (46%) and TC (48%) genotypes exceeded the CC (6%) genotype. For the CAST locus, the MM genotype (83%) was the most frequent, followed by the other genotypes (MN 16% and NN 0.5%) and thus the allelic frequencies were M: 91% and N: 9%. The allele A (66%) of the LEP locus was more frequent than the G allele (34%) and the AA, AG and GG genotypes had frequencies of 43, 47 and 10%, respectively. Only two genotypes (AA 64% and AG 36%) were found in the GH locus with allelic frequencies of 82 and 18% for A and G respectively. In the locus IGF-1, the genotypes AA, GG, and AG presented frequencies of 36, 49 and 15%, respectively, with a greater presence of the G allele (56%) than A (44%). In the analysed population, the GHR and MSTN loci were monomorphic, and only the AA and MM genotypes were found in each. The CAST, GH, and IGF-1 loci showed deviations from HWE. He deficit was found only in the IGF-1 locus. The CAPN, CAST, LEP, GH, and IGF-1 loci were polymorphic with high He and excess heterozygotes, in contrast, the GHR and MST loci were monomorphic. The data presented here could be used in animal breeding programmes.

Expression of growth hormone and growth hormone receptor genes in human eye tissues

In humans, the polygenic growth hormone (GH) locus is located on chromosome 17 and contributes with three types of proteins: pituitary GH which consists of at least two isoforms one of 22 kDa and the other of 20 kDa, placental GH, which also exhibits isoforms, and chorionic somatomammotropin hormone (CSH). While pituitary GH results from the expression of the GH-1 (GH-N) gene, placental GH is produced by the expression of the GH-2 (GH-V) gene and CSH is contributed by expression of the CSH-1 and CSH-2 genes. The location where GH-1 is expressed is the anterior pituitary and the rest of the genes in the locus are expressed in placenta. On the other hand, expression and synthesis of GH in extra-pituitary tissues, including the eye, has been recently described. However, the physiological role of GH in the eye has not yet been elucidated, although a possible neuroprotective role has been hypothesized. Thus, we analyzed GH-1, GH-2, CSH1/2, Pit-1, GHR, GHRH, GHRHR, SST, SSTR1, SSTR2, SSTR3, SSTR4, and SSTR5 to elucidate the expression and regulation of the GH locus in the human eye. Through qPCR analysis, we only found evidence of GH-1 expression in retina, choroid and trabecular meshwork; its transcript turned out to be the same as pituitary GH mRNA found in major species, and no splicing variants were detected. PIT1 was absent in all the ocular tissues implying an independent GH-1 expression mechanism. We found evidence of GHR in the cornea, choroid coat and retina. These results suggest autocrine and/or paracrine regulation, possibly exerted by GHRH and SSTs (since their mRNAs and receptors were found predominantly in retinal, choroidal and corneal tissues) since expression of both molecules was detected in different ocular tissues, as well as in the same tissues where GH-1 expression was confirmed. Our results add solid evidence about the existence of a regulatory local system for GH expression and release in the human eye.

Assessment of growth hormone gene polymorphism effects on reproductive traits in Holstein dairy cattle in Tunisia.

Research to assess the effect of single genes on reproductive traits in bovine species is imperative to elucidate genes' functions and acquire a better perspective of quantitative traits. The present study was undertaken to characterize genetic diversity in the bovine growth hormone (GH) gene in a population of 410 Holstein dairy cows in Tunisia. The analyses were based on single nucleotide polymorphisms, and GH-AluI and GH-MspI detections and genotyping were carried out using the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. Data were analyzed using a mixed linear model with the MIXED procedure to reveal the possible effect of GH genotypes on reproductive traits. The frequency data of AluI(L//V) and MspI(//) alleles were 87.04//12.96 and 70.06//29.94, respectively. The distribution of the frequency of GH genotypes for LL/LV/VV and (/)//(/)//(/) were 77.75//18.59//3.66 and 15.37//29.13//55.50, respectively. The results of the statistical analyses proved that GH-AluI showed a substantial favorable effect on exanimate traits except for the age at first calving; however, only a suggestive effect of GH-MspI on the calving interval (CI) and the days open (DI) was found. The homozygous LL genotype seemed to be advantageous with respect to the CI and the DI compared with LV and VV genotypes. Heterozygous MspI(/) cows tended to have a longer CI and DI than MspI(/) and MspI(/) cows, but the difference was not statistically significant. A significant effect of different GH-AluI–MspI combined genotypes was found on the number of inseminations per conception, the CI and the DI, and the LL/– combined genotype seemed to be associated with better reproductive performance. Based on these results, the LL genotype of the GH locus can be considered to be a favorable genotype for reproductive traits in Holstein dairy cattle, although these findings need to be confirmed by further research before polymorphisms can be used in a marker-assisted selection program.

Single nucleotide polymorphism of the growth hormone (GH) encoding gene in inbred and outbred domestic rabbits

Taking into consideration that the growth hormone (GH) gene in rabbits is a candidate for meat production, understanding the genetic diversity and variation in this locus is of particular relevance. The present study comprised 86 rabbits (Oryctolagus cuniculus) divided into 3 groups: New Zealand White (NZW) outbred rabbits; first-generation inbred rabbits (F<sub>1</sub>) and second-generation inbred rabbits (F<sub>2</sub>). They were analysed by polymerase chain reaction-based restriction fragment length polymorphism method. A 231 bp fragment of the polymorphic site of the GH gene was digested with Bsh1236 restriction enzyme. Single nucleotide polymorphisms for the studied GH locus corresponding to 3 genotypes were detected in the studied rabbit populations: CC, CT and TT. In the synthetic inbred F<sub>1</sub> and F<sub>2</sub> populations, the frequency of the heterozygous genotype CT was 0.696 and 0.609, respectively, while for the homozygous CC genotype the frequency was lower (0.043 and 0.000), and respective values for the homozygous TT genotype were 0.261 and 0.391. This presumed a preponderance of the T allele (0.609 and 0.696) over the C allele (0.391 and 0.304) in these groups. In outbred rabbits, the allele frequencies were 0.613 (allele C) and 0.387 (allele Т); consequently, the frequency of the homozygous CC genotype was higher than that of the homozygous TT genotype (0.300 vs. 0.075). Observed heterozygosity for the GH gene was higher than expected, and the result was therefore a negative inbreeding coefficient (Fis=–0.317 for outbred NZW rabbits; –0.460 for inbred F<sub>1</sub> and –0.438 for inbred F<sub>2</sub>), indicating a sufficient number of heterozygous forms in all studied groups of rabbits. The application of narrow inbreeding by breeding full sibs in the synthetic population did not cause a rapid increase in homozygosity.

Expression of growth hormone gene in the baboon eye

The human growth hormone (GH) locus is comprised by two GH (GH1 and GH2) genes and three chorionic somatomammotropin (CSH1, CSH2 and CSH-L) genes. While GH1 is expressed in the pituitary gland, the rest are expressed in the placenta. However, GH1 is also expressed in several extrapituitary tissues, including the eye. So to understand the role of this hormone in the eye we used the baboon (Papio hamadryas), that like humans has a multigenic GH locus; we set up to investigate the expression and regulation of GH locus in adult and fetal baboon ocular tissues.We searched in baboon ocular tissues the expression of GH1, GH2, CSH1/2, Pit1 (pituitary transcription factor 1), GHR (growth hormone receptor), GHRH (growth hormone releasing hormone), GHRHR (growth hormone releasing hormone receptor), SST (somatostatin), SSTR1 (somatostatin receptor 1), SSTR2 (somatostatin receptor 2), SSTR3 (somatostatin receptor 3), SSTR4 (somatostatin receptor 4), and SSTR5 (somatostatin receptor 5) mRNA transcripts and derived proteins, by qPCR and immunofluorescence assays, respectively. The transcripts found were characterized by cDNA cloning and sequencing, having found only the one belonging to GH1 gene, mainly in the retina/choroid tissues. Through immunofluorescence assays the presence of GH1 and GHR proteins was confirmed in several retinal cell layers. Among the possible neuroendocrine regulators that may control local GH1 expression are GHRH and SST, since their mRNAs and proteins were found mainly in the retina/choroid tissues, as well as their corresponding receptors (GHRH and SSTR1-SSTR5). None of the ocular tissues express Pit1, so gene expression of GH1 in baboon eye could be independent of Pit1. We conclude that to understand the regulation of GH in the human eye, the baboon offers a very good experimental model.

The Associations of GH and GHR Genes with Carcass Components in Indonesian Kampung and Broiler Chicken Cross

The chicken growth hormone (GH) and its receptor (growth hormone receptor, GHR) play important roles in chicken performances due to their crucial functions in growth. The variations of GH and GHR genes were then thought to be associated with the variations of the performances. This experiment was designed to identify the g.2248G>A GH and the g.565G>A GHR loci polymorphisms and to evaluate their associations with carcass components in Kampung and broiler chicken cross. A total of 215 chickens including 4 chicken populations (Kampung, Cobb broiler, F1, and F2 Kampung x broiler chicken cross) were screened to identify polymorphism using PCR-RFLP technique with EcoRV and Eco72I restriction enzyme for GH and GHR loci, respectively. The carcass components were recorded at 26 weeks of age on F2 Kampung and broiler chicken cross (42 chickens) for association study. Both the g.2248G>A GH and the g.565G>A GHR loci were polymorphic with two alleles (G and A) and three genotypes (GG, AG, and AA). The GG genotype and the G allele of GH locus were predominant in all chicken populations. While in GHR locus, the AA genotype and the A allele were found to be higher in all chicken populations. The association study showed that the g.565G>A GHR locus polymorphism had significant effect on carcass components, including live weight, carcass weight, breast weight, thighs weight, breast muscle weight, and thighs muscle weight. There was no significant association was found between the g.2248G>A GH genotype and carcass components. It could be concluded that the g.2248G>A GH and the g.565G>A GHR loci were polymorphic in Kampung and broiler chicken cross and the g.565G>A GHR locus was accosiated with carcass components. This g.565G>A GHR SNP might be an important candidate marker for chicken growth and muscle mass improvement.

Structure and evolution of the gorilla and orangutan growth hormone loci.

In primates, the unigenic growth hormone (GH) locus of prosimians expressed primarily in the anterior pituitary, evolved by gene duplications, independently in New World Monkeys (NWM) and Old World Monkeys (OWMs)/apes, to give complex clusters of genes expressed in the pituitary and placenta. In human and chimpanzee, the GH locus comprises five genes, GH-N being expressed as pituitary GH, whereas GH-V (placental GH) and CSHs (chorionic somatomammotropins) are expressed (in human and probably chimpanzee) in the placenta; the CSHs comprise CSH-A, CSH-B and the aberrant CSH-L (possibly a pseudogene) in human, and CSH-A1, CSH-A2 and CSH-B in chimpanzee. Here, the GH locus in two additional great apes, gorilla (Gorilla gorilla gorilla) and orangutan (Pongo abelii), is shown to contain six and four GH-like genes, respectively. The gorilla locus possesses six potentially expressed genes, gGH-N, gGH-V and four gCSHs, whereas the orangutan locus has just three functional genes, oGH-N, oGH-V and oCSH-B, plus a pseudogene, oCSH-L. Analysis of regulatory sequences, including promoter, enhancer and P-elements, shows significant variation; in particular the proximal Pit-1 element of GH-V genes differs markedly from that of other genes in the cluster. Phylogenetic analysis shows that the initial gene duplication led to distinct GH-like and CSH-like genes and that a second duplication provided separate GH-N and GH-V. However, evolution of the CSH-like genes remains unclear. Rapid adaptive evolution gave rise to the distinct CSHs, after the first duplication, and to GH-V after the second duplication. Analysis of transcriptomic databases derived from gorilla tissues establishes that the gGH-N, gGH-V and several gCSH genes are expressed, but the significance of the many CSH genes in gorilla remains unclear.

HERITABILITIES OF BODY SIZE BY GROWTH HORMONE (GH-Msp1) GENOTYPES USING PCR-RFLP IN ONGOLE GRADE CATTLE

Genotypic performance in term of heritability as the crucial factor of animal economical traits for body size inheritance had not been fully studied in Ongole-grade cattle. The objectives of this research were to define the heritability values of live weight, chest girth and body length in Ongole-crossbred cattle. Total of 37 blood samples were collected from parental cows and 2 blood samples from parental Ongole breed bulls. All blood samples were screened for the presence of growth hormone ( GH ) locus using PCR-RFLP method involving restricted enzyme Msp1 on agarose-gel (1.2%). Data were analyzed using statistical program in Excel XP. Results showed that the phenotypic estimation average of Ongole grade cattle population of live weight, chest girth and body length were 445.41 ± 45.95 kg, 175.35 ± 4.11 cm, and 139.70 ± 5.73cm, respectively. The heritability values of animal live weight, chest girth and body length in this study were 0.24, 0.003, and 0.41, respectively. These heritability values of animal live weight and body length would be categorized as moderate to high genotypic performance values, while the heritability of animal chest girth was included in low heritability standard of the animal economical trait performance.

Determination of the AluI polymorphism effect of bovine growth hormone gene on carcass traits in Zavot cattle with analysis of covariance

The aim of this study was to determine the genotypes of bovine growth hormone and to investigate their association with daily weight gain in the Zavot cattle breed. The bovine growth hormone (GH) genotypes of 45 male Zavot cattle were identified by PCR-RFLP. The effect of the bovine growth hormone gene on daily weight gain was estimated from a dataset obtained from cattle up to the age of 9 months. The mean differences of carcass weight or carcass yield among the genotype groups were assessed by analysis of covariance (ANCOVA). Two alleles for bovine growth hormone locus, L and V, were observed in Zavot cattle. The frequency of the L and V alleles was found to be 0.767 and 0.233, respectively. The genotype effect on live weight was not found to be significant (P > 0.05). The difference between the means of genotype groups was not found to be significant for carcass weight and carcass yield (P > 0.05). Further research on the association between bovine growth hormone gene polymorphism and yield traits in different cattle breeds should be conducted.

Tissue specific CTCF occupancy and boundary function at the human growth hormone locus

The robust and tissue-specific activation of the human growth hormone (hGH) gene cluster in the pituitary and placenta constitutes an informative model for analysis of gene regulation. The five-gene hGH cluster is regulated by two partially overlapping sets of DNase I hypersensitive sites (HSs) that constitute the pituitary (HSI, II, III and V) and placental (HSIII, IV, and V) locus control regions (LCRs). The single placenta-specific LCR component, HSIV, is located at −30 kb to the cluster. Here we generate a series of hGH/BAC transgenes specifically modified to identify structural features of the hGH locus required for its appropriate placental expression. We find that placental specificity is dependent on the overall multigene configuration of the cluster whereas the distance between the cluster and its LCR impacts the level of placental expression. We further observe that a major function of the placental hGH LCR is to insulate the transgene locus from site-of-integration effects. This insulation activity is linked to placenta-specific occupancy of the chromatin architectural protein, CTCF, at HSIV. These data reveal a remarkable combination of structural configurations and regulatory determinants that must work in concert to insure robust and tightly controlled expression from a complex multigene locus.

Distinct Chromatin Configurations Regulate the Initiation and the Maintenance of hGH Gene Expression

For many mammalian genes, initiation of transcription during embryonic development must be subsequently sustained over extensive periods of adult life. It remains unclear whether maintenance of gene expression reflects the same set of pathways as are involved in initial gene activation. The human pituitary growth hormone (hGH-N) locus is activated in the differentiating somatotrope midway through embryogenesis by a multicomponent locus control region (LCR). DNase I-hypersensitive site I (HSI) of the LCR is essential to full developmental activation of the hGH-N locus. Here we demonstrate that conditional deletion of HSI from the active hGH locus in the adult pituitary effectively silences hGH-N expression. Analyses of chromatin structure and locus positioning demonstrate that a specific subset of the HSI functions active in the embryo retain their HSI dependence in the adult pituitary. These functions sustain engagement of the hGH locus with polymerase II (Pol II) factories, histone acetylation at the hGH-N promoter, and looping of the LCR to its target promoter. These data reveal that HSI is essential to both the maintenance and the initiation phases of gene expression. These observations contribute to our mechanistic understanding of how stable patterns of mammalian gene expression are established in a terminally differentiated cell.

Polymorphisms of Growth Hormone Gene in a Native Chicken Population: Association with Egg Production

Abstract A total of 142 chicken blood samples were collected and a specific primer set was used to amplify a fragment of growth hormone locus using PCR. PCR products were digested with SacI and MspI restriction endonucleases. The amplified fragment digested with SacI enzyme revealed two “+” (wild type) and “-” (normal type) alleles with the frequency of 0.898 and 0.102, respectively. The amplified fragment digested with MspI enzyme revealed three A, B and C alleles with the frequency of 0.599, 0.102, and 0.299, respectively. Frequencies of +/+, +/- and -/- were 0.817, 0.162, and 0.021, respectively, and those of AA, AB, AC, BB, BC, and CC were 0.338, 0.113, 0.409, 0.007, 0.070, and 0.063, respectively, in the studied population. The results of 2 and likelihood ratio tests showed that this population was at Hardy-Weinberg equilibrium with respect to the marker locus. Marker-trait association analysis revealed statistically significant differences between “SacI-RFLP” genotypes for egg production and rate of laying eggs. The relationship between the molecular marker and these traits can be useful to improve the chicken breeding programmes.

DNase I Hypersensitive Site II of the Human Growth Hormone Locus Control Region Mediates an Essential and Distinct Long-range Enhancer Function

Locus control regions (LCRs) comprise sets of DNA elements capable of establishing autonomous chromatin domains that support robust and physiologically appropriate expression of target genes, often working over extensive distances. Human growth hormone (hGH-N) expression in the pituitary is under the regulation of a well characterized LCR containing four DNase I hypersensitive sites (HSs). The two pituitary-specific HS, HSI and HSII, are located 14.5 and 15.5 kb 5' to the hGH-N promoter. HSI is essential for activation of hGH-N during pituitary development and for sustaining robust activity in the adult. To determine whether the closely linked HSII has a role in hGH-N expression, it was deleted from a previously validated hGH/P1 transgene. Analysis of three independent hGH/P1(ΔHSII) transgenic mouse lines revealed that this deletion had no adverse effect on the formation of HSI, yet resulted in a substantial loss (70%) in hGH-N mRNA expression. This loss of expression was accompanied by a corresponding reduction in recruitment of the pituitary-specific transcription factor Pit-1 to the hGH-N promoter and a selective decrease in promoter occupancy of the elongation-linked isoform of RNA polymerase II. Sufficiency of HSI and HSII in LCR activity was explored by establishing two additional sets of mouse transgenic lines in which DNA segments containing these HS were positioned within the λ phage genome. In this "neutral" DNA context, HSII was required for the recruitment of HAT activity. These data establish HSII as a nonredundant component of the hGH LCR essential for establishment of robust levels of hGH-N gene expression.