Flanking Region Of Prolactin Gene Research Articles

Genetic polymorphism at 5′ flanking region of the prolactin gene and its effect on egg quality traits in naked neck chickens

A study was conducted to explore polymorphism at 5′ flanking region of prolactin gene and to estimate the effect of polymorphism on egg quality traits in naked neck chickens. Single-stranded conformation polymorphism technique was employed to detect polymorphism in the gene. Three genotypes (AA, AB and AC) were observed in naked neck chicken of which the predominant genotype was AB. The predominant and least frequent alleles in the population were A and C, respectively. The sequence alignment study revealed a number of nucleotide substitutions at different positions of the alleles. Genotypes showed significant effect (P < 0.05) on Haugh unit and yolk index where birds with AB and AC genotypes were the best performers. In conclusion, it may be inferred that 5′ flanking region of prolactin gene was polymorphic with having significant effect on some egg quality traits in the naked neck chicken.

DNA polymorphism of 5′ flanking region of prolactin gene and its association with litter size in sheep

A single nucleotide polymorphism of 5' flanking region of the prolactin gene was investigated in both high prolificacy breeds (Small Tail Han and Hu sheep) and low prolificacy breeds (Dorset and Suffolk sheep) using polymerase chain reaction (PCR)-single strand conformation polymorphism (SSCP). The results indicated that two genotypes (AA and AB) were detected in Small Tail Han sheep (n = 239), only one genotype (AA) was detected in Hu (n = 40), Dorset (n = 50) and Suffolk sheep (n = 39). The mutant homozygous genotype (BB) was not detected in four sheep breeds. In Small Tail Han sheep (n = 239), the frequency of genotypes AA and AB was 0.91 and 0.09, the frequency of the A and B alleles was 0.95 and 0.05, respectively. The fitness tests showed that the Small Tail Han sheep population was in Hardy-Weinberg equilibrium. Sequencing revealed a mutation (G-->T) at the position 63 bp of the 5' flanking region of prolactin gene in AB genotype compared with AA genotype in Small Tail Han sheep. The Small Tail Han ewes with AB genotype had 0.83 (p < 0.05) lambs more than those with AA genotype. These results preliminarily showed that the prolactin locus is either a major gene that influences the high prolificacy in Small Tail Han sheep or is in close linkage with such a gene.

Cis-acting negative regulatory element of prolactin gene.

The prolactin (PRL) gene in clonal strains of rat pituitary tumor cells in culture (GH cells) exhibit several regulatory responses, similar to the ones observed in rat pituitary gland. A comparative analysis of regulation of PRL gene expression in PRL-producing (PRL+) and PRL-nonproducing (PRL-) GH cells was conducted by monitoring the PRL promoter driven transient expression of chloramphenicol acetyltransferase (CAT) gene in GH4C1 (PRL+) and GH12C1 (PRL-) cells. The PRL promoter activity was drastically inhibited only in PRL-nonproducing cells (PRL-) and not in PRL producing cells (PRL+) when a 80-base pair (bp) DNA sequence from 5'-flanking region of PRL gene (located between -330 and -250 bp) was included in the PRL-CAT fusion gene constructs. Furthermore, a DNA/protein interaction involving this 80-bp DNA sequence and a 60-kDa nuclear protein was detected only in PRL- cells but not in PRL+ GH cells. These results suggested that the strain-specific suppression of PRL gene in PRL-, GH12C1 cells was mediated via interaction of a cis-acting negative regulatory element with a negative regulatory trans-acting factor in these cells. The negative regulatory element within the AT-rich 80-bp DNA sequence was mapped immediately adjacent to the site of interaction of trans-activators of PRL gene.

Detection of two chromatin proteins which bind specifically to the 5'-flanking region of the rat prolactin gene.

We employed a protein gel blotting procedure to search for nuclear proteins from rat pituitary cells that bind preferentially to the 5'-flanking region of the rat prolactin gene. By gel blots of chromatin proteins from GH3 rat pituitary tumor cells with a 32P-labeled prolactin genomic clone, we detected two major binding proteins with molecular weights of approximately 44,000 and 48,000, designated NP44 and NP48, respectively. Both NP44 and NP48 are minor chromatin proteins which are extracted at low salt concentrations (0.4 M NaCl) and exhibit a range of slightly acidic isoelectric variants. NP44 and NP48 were detected at similar levels in chromatin extracts of GH3 cells, the prolactin-negative GC cell variant of the GH3 cells, and normal rat pituitary tissue. Considerably lower levels of these two proteins were found in chromatin extracts from rat liver and rat C6 glial cells. NP44 and NP48 exhibit DNA sequence specificity, as evidenced by their strong binding to the upstream flanking region of the prolactin gene, but only very weak binding to plasmid DNA, rat prolactin or growth hormone cDNAs, or upstream flanking regions of two other rat genes. By analyzing subclones of a rat prolactin genomic clone, we established that NP44 and NP48 bind to at least two sites, which are located between 0.4 and 2.0 kilobases (region I) and between 2.0 and 4.8 kilobases (region II) upstream of the transcription initiation site. These findings are discussed in the context of a possible functional association between the strong binding of NP44 and NP48 to the prolactin 5'-flanking region and pituitary-specific expression of the prolactin gene.

Detection of two chromatin proteins which bind specifically to the 5'-flanking region of the rat prolactin gene.

We employed a protein gel blotting procedure to search for nuclear proteins from rat pituitary cells that bind preferentially to the 5'-flanking region of the rat prolactin gene. By gel blots of chromatin proteins from GH3 rat pituitary tumor cells with a 32P-labeled prolactin genomic clone, we detected two major binding proteins with molecular weights of approximately 44,000 and 48,000, designated NP44 and NP48, respectively. Both NP44 and NP48 are minor chromatin proteins which are extracted at low salt concentrations (0.4 M NaCl) and exhibit a range of slightly acidic isoelectric variants. NP44 and NP48 were detected at similar levels in chromatin extracts of GH3 cells, the prolactin-negative GC cell variant of the GH3 cells, and normal rat pituitary tissue. Considerably lower levels of these two proteins were found in chromatin extracts from rat liver and rat C6 glial cells. NP44 and NP48 exhibit DNA sequence specificity, as evidenced by their strong binding to the upstream flanking region of the prolactin gene, but only very weak binding to plasmid DNA, rat prolactin or growth hormone cDNAs, or upstream flanking regions of two other rat genes. By analyzing subclones of a rat prolactin genomic clone, we established that NP44 and NP48 bind to at least two sites, which are located between 0.4 and 2.0 kilobases (region I) and between 2.0 and 4.8 kilobases (region II) upstream of the transcription initiation site. These findings are discussed in the context of a possible functional association between the strong binding of NP44 and NP48 to the prolactin 5'-flanking region and pituitary-specific expression of the prolactin gene.

Selective binding of the estradiol receptor to a region at least one kilobase upstream from the rat prolactin gene.

The interaction of the estradiol receptor with cloned DNA fragments from the prolactin gene was investigated using a competitive binding assay. A DNA fragment from the 5'-flanking region of the rat prolactin gene was able to bind the estradiol receptor selectively. DNA fragments representing most of the remaining 10 kb of the prolactin gene showed little or no selective binding to the estradiol receptor. The fragment from the 5'-flanking region which selectively binds the receptor is located between 1.2 and 2.0 kb upstream from the transcription initiation site rather than in a more proximal position as has been observed for the interaction of several other steroid hormone receptors with specific genes. Nucleotide sequence analysis demonstrated that this region of DNA contains two large segments of alternating purine-pyrimidine sequence. One of the alternating purine-pyrimidine regions is very large, containing more than 160 nucleotides of almost perfect poly(dG-dT). Further studies will be required to determine if the receptor interacts specifically with these interesting sequences. However, the ability of the estradiol receptor to bind to an upstream 5'-flanking region of the prolactin gene may be part of the mechanism which allows the receptor to stimulate the transcription of this gene selectively.