Prolactin Receptor Gene Research Articles

Gene Editing Cattle for Enhancing Heat Tolerance: A Welfare Review of the “PRLR-SLICK Cattle” Case

In March 2022 the US Food and Drug Administration (FDA) published a risk assessment of a recent animal gene editing proposal submitted by Acceligen™. The proposal concerned the possibility of changing the cattle genome to obtain a slicker, shorter hair coat. Using CRISPR-Cas9 it was possible to introduce an intentional genomic alteration (IGA) to the prolactin receptor gene (PRLR), thereby producing PRLR-SLICK cattle. The goal was to diminish heat stress in the cattle by enhancing their heat-tolerance. With regard to unintended alterations (i.e., off-target effects), the FDA stated that the IGA posed a low, but still present, risk to animal safety. The aim of this article is to present some initial insights into the welfare issues raised by PRLR-SLICK cattle by addressing the question: Do SLICK cattle have better welfare than non-SLICK cattle when exposed to heat stress? Two potential welfare concerns are examined. The first is pleiotropy, an issue that arises when one gene affects multiple traits. Given the pleiotropic nature of prolactin, it has been suggested that the IGA for SLICK cattle may also affect their hepatic and other functions. The second concern relates not primarily to direct effects on cattle health, but rather to the indirect risk that this more heat-tolerant animal would just be used in the livestock sector under farming conditions that are such that the net welfare improvement would be non-existent.

Consequences of gene editing of PRLR on thermotolerance, growth, and male reproduction in cattle.

Global warming is a major challenge to the sustainable and humane production of food because of the increased risk of livestock to heat stress. Here, the example of the prolactin receptor (PRLR) gene is used to demonstrate how gene editing can increase the resistance of cattle to heat stress by the introduction of mutations conferring thermotolerance. Several cattle populations in South and Central America possess natural mutations in PRLR that result in affected animals having short hair and being thermotolerant. CRISPR/Cas9 technology was used to introduce variants of PRLR in two thermosensitive breeds of cattle - Angus and Jersey. Gene-edited animals exhibited superior ability to regulate vaginal temperature (heifers) and rectal temperature (bulls) compared to animals that were not gene-edited. Moreover, gene-edited animals exhibited superior growth characteristics and had larger scrotal circumference. There was no evidence for deleterious effects of the mutation on carcass characteristics or male reproductive function. These results indicate the potential for reducing heat stress in relevant environments to enhance cattle productivity.

Significance of Duodenal Prolactin Receptor Modulation by Calcium and Vitamin D in Sulpiride-Induced Hyperprolactinemia.

Background and Objectives: Hyperprolactinemia, as a potential side-effect of some antipsychotic medications, is associated with decreased bone density and an increased risk of fractures. This study investigates whether calcium and vitamin D supplementation affects prolactin receptor (Prlr) gene expression in the duodenum, vertebrae, and kidneys of female rats with sulpiride-induced hyperprolactinemia. Materials and Methods: Twenty-one-week-old female Wistar rats were assigned to three groups: Group S consisted of ten rats who received sulpiride injections (10 mg/kg) twice daily for 6 weeks; Group D (10 rats) received daily supplementation of 50 mg calcium and 500 IU vitamin D along with sulpiride for the last 3 weeks; and Group C consisting of seven age-matched nulliparous rats serving as a control group. Real-time PCR was used to assess Prlr gene expression in the duodenum, vertebrae, and kidneys. Results: In Group S, Prlr gene expression was notably decreased in the duodenum (p < 0.01) but elevated in the vertebrae and kidneys compared to Group C. Conversely, Group D exhibited significantly increased Prlr expression in the duodenum (p < 0.01) alongside elevated expression in the vertebrae and kidneys. Conclusions: In sulpiride-induced hyperprolactinemia, decreased Prlr gene expression in the duodenum may lead to reduced intestinal calcium absorption. Consequently, prolactin may draw calcium from the skeletal system to maintain calcium balance, facilitated by increased Prlr gene expression in the vertebrae. However, vitamin D supplementation in sulpiride-induced hyperprolactinemia notably enhances Prlr gene expression in the duodenum, potentially ameliorating intestinal calcium absorption and mitigating adverse effects on bone health.

Comprehensive in silico analysis of prolactin receptor (PRLR) gene nonsynonymous single nucleotide polymorphisms (nsSNPs) reveals multifaceted impact on protein structure, function, and interactions

This study delves into the functional and structural implications of non-synonymous single nucleotide polymorphisms (nsSNPs) within the Prolactin Receptor (PRLR) gene. Thirteen deleterious nsSNPs were identified through bioinformatics tools, with SIFT predicting 168 out of 395 nsSNPs as detrimental, exhibiting tolerance index (TI) scores ranging from 0 to 0.05. Polyphen2 assigned likelihood scores >0.99 to all 13 nsSNPs, indicating high probability of harm, while Panther scores classified most nsSNPs as ‘probably damaging’, with specific mutations like W218R scoring 0.74, suggesting a higher impact. Stability analysis using DDG I-Mutant and DDG Mupro consistently predicted decreased stability for all mutations, with CUPSAT indicating mutations like V125G and W218R significantly decreasing stability. Structural analysis through DynaMut predicted destabilization for all mutations except L196I and L292H. MutPred2 highlighted structural alterations for all nsSNPs except L196I, L293V, R315W, and S353N. Domain analysis revealed key mutations within essential functional domains, with five nsSNPs located within Fibronectin type-III domains. Bayesian analysis through ConSurf identified 9 critical residues, with 11 nsSNPs exhibiting notably high conservation. STRING analysis unveiled a complex interaction network, indicating involvement in vital biological processes like lactation. Molecular dynamics (MD) simulations, spanning 100 nanoseconds, elucidated structural dynamics induced by detrimental missense SNPs. Post-translational modification (PTM) analysis identified specific mutations, such as R351, involved in methylation, while S353 was implicated in phosphorylation and glycosylation. These findings offer comprehensive insights into the molecular and phenotypic effects of deleterious nsSNPs in the PRLR gene, crucial for selective breeding. Communicated by Ramaswamy H. Sarma

Thermotolerance capabilities, blood metabolomics, and mammary gland hemodynamics and transcriptomic profiles of slick-haired Holstein cattle during mid lactation in Puerto Rico

Holstein cattle carrying a prolactin receptor gene mutation (SLICK) exhibit short and sleek hair coats (short-haired Holstein [SLK]) enhancing thermotolerance and productivity compared with wild type-haired Holstein (WT) under tropical conditions. The objectives were to unravel the physiological and molecular mechanisms that confer an advantage to this slick genotype in Puerto Rico and determine potential correlations between metabolites and physiological variables. At 160 ± 3 DIM we compared vaginal temperatures (VT) and voluntary solar radiation exposure (VSRE) during 48 h between 9 SLK and 9 WT Holsteins, whereas a subsample of 7 SLK and 7 WT were used to assess udder skin temperature, mammary gland hemodynamics and transcriptomics, and blood plasma untargeted metabolomics at a single time point. The SLK cattle showed lower VT throughout the day and greater VSRE at 1000 h and 1100 h compared with their WT counterparts. Total mammary blood flow (MBF) was greater in SLK Holsteins compared with WT. The metabolite 9-nitrooctadecenoic acid was identified as a potential biomarker for MBF; moreover, SLK cattle had greater amounts of this metabolite in their plasma. Prostaglandin D2 synthase (PTGS) was upregulated in the slick mammary gland, while plasma prostaglandin D2 was positively correlated with milk yield and increased in SLK Holsteins compared with WT. Interestingly, the arachidonic acid metabolism pathway was enriched in the mammary gland transcriptome and perturbed in the blood metabolome in the SLK Holsteins. In conclusion, SLK Holsteins exhibited lower body temperatures, greater VSRE, enhanced blood supply to the mammary gland, and alterations in genes and metabolites involved in arachidonic acid metabolism at the mammary gland and blood plasma. The usage of the SLK Holstein cattle genetics in dairy operations could be a feasible alternative to mitigate the adverse consequences of heat stress.

Standardized mixture of Pluchea indica and Sauropus androgynus extract stimulates the gene expression associated with lactogenesis in rats

Pluchea indica (P. indica) and Sauropus androgynus (S. androgynus) are medicinal plants empirically used as an herbal galactogogue, but their mechanism of action has not been elucidated at the molecular level in the form of a mixed extract. This research aimed to standardize and investigate the combined effects of P. indica leaf extract (EPI) and S. androgynus leaf extract (ESA) on the expressions of prolactin (PRL), PRL receptor (PRLR), oxytocin (OXT), and OXT receptor (OXTR) genes in pituitary and mammary glands of lactating rats. The extracts were prepared by maceration in 70% ethanol. Standardization was done by examining non-specific parameters and determining total flavonoids using spectrophotometric analysis. Twenty-four lactating rats were divided into six groups. Group I: CMC 0.5% as control, II: EPI 500 mg/kgBW, III: ESA 125 mg/kgBW, and IV–VI were given combination formula EPI-ESA 1: 125+31.25, EPI-ESA 2: 250+62.5, and EPI-ESA 3: 500+125 mg/kgBW (orally, once a day) from the 2nd–15th day after parturition. On the 16th day, experimental animals were euthanized for tissue sampling, and gene expression was measured using real-time reverse transcription-quantitative polymerase chain reaction (RT-qPCR). The study of the non-specific parameters revealed water content, total ash, and acid-insoluble ash met criteria based on the Indonesian Herbal Pharmacopoeia. Evaluation of total flavonoids yielded 7.33% ± 0.00% and 3.84% ± 0.01% w/w for EPI and ESA, respectively. Test results showed balanced induction of all target genes was achieved when subjects received EPI-ESA 2. We concluded that EPI-ESA can potentially support lactogenesis by stimulating the expressions of PRL, PRLR, OXT, and OXTR genes.

76 Transcriptional profiles and signaling pathways associated with the SLICK1 allele of the prolactin receptor gene in Holstein cattle

76 Transcriptional profiles and signaling pathways associated with the SLICK1 allele of the prolactin receptor gene in Holstein cattle

The prolactin receptor gene (PRLR) is linked and associated with the risk of polycystic ovarian syndrome

The prolactin receptor gene (PRLR) may contribute to polycystic ovarian syndrome (PCOS) since it plays important roles in physiological ovarian functions. PRLR-knockout mice have irregular cycles and subfertility and variants in or around the PRLR gene were associated in humans with female testosterone levels and recurrent miscarriage. We tested 40 variants in the PRLR gene in 212 Italian families phenotyped by type 2 diabetes (T2D) and PCOS and found two intronic PRLR-variants (rs13436213 and rs1604428) significantly linked to and/or associated with the risk of PCOS. This is the first study to report PRLR as a novel risk gene in PCOS. Functional studies are needed to confirm these results.

PSXIV-17 Examining Mammary Gland Transcriptomic Profiles Using Rna-Seq in Slick and Wild-Type Haired Holstein Cattle Under Tropical Conditions

Abstract In lactating dairy cows, heat stress reduces feed intake, alters the metabolism of the animal, and compromises milk production. Holstein cattle carrying the prolactin receptor gene mutation (SLICK) have been shown to have superior performance under tropical conditions by exhibiting increased milk yield compared with wild-type haired Holstein cattle (WT). Recent research indicated that SLICK cattle have greater mammary gland blood flow and lower vaginal temperatures, while maintaining prolonged voluntary exposure to direct solar radiation than their WT counterparts. Nevertheless, differences in molecular mechanisms between genotypes at the mammary gland level have been unexplored. The objective of the current study was to investigate possible differences in the mammary gland transcriptome from SLICK (n = 7) and WT (n = 6) Puerto Rican Holstein cattle. At 160 ± 3 days in milk (DIM), ultrasound guided mammary biopsies were collected from the left rear quarter of each animal and samples were snap frozen. Unstranded RNA-seq libraries were created from the frozen mammary tissue and sequenced using Illumina Novaseq 2x150bp. Quantification of expression was done by salmon [v1.9.0] using the Bos taurus (ARS UCD1.3) transcriptome database. Testing for significant differential expression (DE) was done using a pipeline consisting of the R packages tximport [v1.26.1] and edgeR [v3.40.2]. The fry method from edgeR was used to do gene set enrichment analysis (GSEA) for Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways and Gene Ontology (GO) terms. A single gene (GeneID:513329, major allergen Equ c 1 isoform X2) showed significant (B.H. adjusted P-value ≤ 0.05) differential expression. Aligning the protein sequence for this gene using the BLASTp against the non-redundant (NR) database revealed high similarity to two pheromones, salivary lipocalin 2 and salivary lipocalin-like. Both the KEGG pathway and GO term GSEA showed no statistically significant enrichment at B.H. adjusted P-value ≤ 0.05; however, using a less stringent criteria with a normal P-value ≤ 0.05, multiple relevant pathways were identified. For example, the KEGG pathway analysis revealed that the arachidonic acid metabolism pathway (bta00590) was enriched, and the tight junction function pathway (bta4530) was depleted in the SLICK Holstein samples. Among the GO analysis, genes were found to be downregulated in response to cold (GO:0009409), hair follicle development (GO:0001942), and vasoconstriction (GO:0042310), whereas positive regulation of oxytocin production (GO:0140668), and defense response to Gram-positive bacterium (GO:0050830) were upregulated. In conclusion, differences in gene expression were limited, but after enrichment interesting pathways were revealed and deserve further exploration to aid in elucidating different molecular mechanisms between SLICK and WT cattle.

Genes of Hormone Receptors Affecting Egg Productivity and Reproductive Qualities of Chickens

The egg production of chickens is a complex polygenic type of inheritance and is controlled by many genes. The laying performance of chickens is the result of a complex process regulated by the hypothalamic-pituitary-gonadal system. This review summarizes the information on the effect of polymorphisms of follicle stimulating hormone (FSHR), luteinizing hormone (LHCGR), progesterone (PR) and prolactin (PRLR) receptor genes on the egg production and reproductive performance of chickens. The data presented show that the polymorphisms of these genes are promising for use in breeding programs to improve egg production and reproductive performance of chickens.

FRI303 PRL-R Variants Are Not Only Associated With Prolactinomas But Also With Dopamine Agonist Resistance

Abstract Disclosure: A.R. Moreira: None. E.B. Trarbach: None. A. Glezer: None. G.A. Maciel: None. C.B. Bueno: None. A.S. Monteiro: None. M. Padula: None. I.P. Grande: None. M.D. Bronstein: None. Introduction: Knockout prolactin receptor gene (PRL-R) mice are animal models for prolactinomas and PRL acts via autocrine/paracrine inhibiting lactotroph proliferation. Recent studies identified variants in PRL-R gene in patients with prolactinoma and two of them Glu400(376)Gln/p.Asn516(492)Ile, were more common in patients compared to controls. Functional analysis of the p.Asn516(492)Ile variant showed an increase in Akt expression and cell proliferation induced by PRL. However, the association between PRL-R variants and response to cabergoline (CAB) treatment has not been investigated. Objective: This study aims to analyze the PRL-R gene in an extensive cohort of patients with sporadic prolactinomas and evaluate its association with clinical, laboratorial and tumor imaging characteristics, as well as CAB treatment resistance. Methods: DNA was extracted from peripheral blood samples of 178 patients with prolactinoma and 146 were classified according to cabergoline response. The PRL-R gene was analyzed by PCR techniques and automated Sanger-type sequencing. Four in silico tools were used for analysis. Results: We found six PRL-R variants: p.Ile100(76)Val (n=18), p.Ile170(146)Leu (n=6), p.Glu400(376)Gln/p.Asn516(492)Ile (n=3), p.Glu470Asp (n =1) and p.Ala591Pro (n=1), the last two described for the first time in prolactinomas´ patients. The variants p.Glu400(376)Gln/p.Asn516(492)Ile and p.Ala591Pro were more frequent amongst patients compared to genomic databases and the p.Asn516(492)Ile showed pathogenic potential using in silico analysis. PRL-R variants were associated with male sex (P=0.015), higher serum PRL levels (P=0.007), larger tumors (P=0.001), higher frequency of macroadenomas (P=0.031) and cabergoline resistance (P&amp;lt;0.001). Considering together only the PRL-R variants classified as pathogenic/probably pathogenic (p.Ile170(146)Leu, p.Glu400(376)Gln/Asn516(492) and p.Ala591Pro) versus the benign variants (p.Ile100(76)Val, p.Glu470Asp and wild type) we also observed a higher levels of serum PRL (P=0.008), largest tumor diameter (P=0.028) and higher frequency of CAB resistance (P=0.048).Conclusion: Our study showed a higher frequency of the PRL-R variants p.Glu400(376)Gln/p.Asn516(492)Ile, and p.Ala591Pro in patients with prolactinomas when compared to Brazilian and international databases, suggesting that these variants could play a role in the prolactinoma tumorigenesis. Patients with PRL-R variants presented male predominance, higher serum PRL levels, larger tumor size at diagnosis and higher frequency of CAB resistance. More studies evaluating germinative and somatic PRL-R variants are needed to clarify the PRL/PRL-R system role in the tumorigenesis and progression of prolactinomas and their interesting association with CAB resistance. Presentation: Friday, June 16, 2023

A novel signaling transduction pathway of melatonin on lactose synthesis in cows via melatonin receptor 1 (MT1) and prolactin receptor (PRLR).

In the current study, we explored the relationship between melatonin and lactose synthesis in in vivo and in vitro conditions. We found that long-term melatonin feeding to the dairy cows significantly reduced the milk lactose content in a dose dependent manner. This lactose reduction was not associated with a negative energy balance, since melatonin treatment did not alter the fat, glucose, or protein metabolisms of the cows. To identify the potential molecular mechanisms, the cow's mammary epithelial cells were cultured for gene expression analysis. The results showed that the effect of melatonin on lactose reduction was mediated by its receptor MT1. MT1 activation downregulated the mRNA expression of the prolactin receptor gene (PRLR), which then suppressed the gene expression of SLC35B1. SLC35B1 is a galactose transporter and is responsible for the transportation of galactose to Golgi apparatus for lactose synthesis. Its suppression reduced the lactose synthesis and the milk lactose content. The discovery of this signal transduction pathway of melatonin on lactose synthesis provides a novel aspect of melatonin's effect on carbohydrate metabolism in cows and maybe also in other mammals, including humans.

Analysis of polymorphic variants of MSTN, CAST, PRLR genes associations with economically useful qualities of vyatka breed horses

The relationship of polymorphic variants of MSTN, CAST, PRLR genes with working qualities and body types of Vyatka horses was studied. The purpose of the research is to assess the genetic and breeding potential of the Vyatka breed horses, as well as to study the relationship between the MSTN, CAST, PRLR genotypes and the economically useful qualities of horses. The method of DNA extraction from Vyatka horse hair follicles using ExtraGene DNA Prep. was applied. When scanning the mutations in the loci MSTN (n=43), CAST (n=41) and PRLR (n=41), DNA amplification the method of allele-specific PCR was performed. The frequency of alleles and genotypes was calculated using MS Excel 10. As a result of research, the dependence of Vyatka horses working qualities on the frequency of occurrence of myostatin MSTN alleles (g.66493737 T&gt;C) was noted. The horses with a higher occurrence of the MSTN/C allele have more productive movements than the individuals with the T/T genotype typical for aborigines. Vyatka horses with the T/T genotype are more versatile, and also show better results in sledding, while the horses with the T/C genotype are better under saddle. The horses with the T/T genotype have the highest bony index but the lowest massiveness index. The MSTN T/T (0.581) homozygous genotype for the "wild" allele predominates in Vyatka horses. A relation ship between body types and the calpastatin gene (CAST) has been revealed for the first time in horse breeding. The horses with the G/A genotype turned out to be the most massive and bony, the horses with the A/A genotype were lighter, the highest frequency of occurrence of the CAST G/A genotype (0.463) was noted, the CAST G/G genotype is rare in the breed (0.171). A relationship between the frequency of occurrence of prolactin receptor genes (PRLR) and body types of horses was not found. The frequency of occurrence of PRLR C/C (0.366) and PRLR G/C (0.390) genotypes is approximately identical, the PRLR G/G genotype is less common (0.244). The study of genes associated with economically useful qualities in all breeding stallions will enable to conduct more efficient breeding, using the desired genotypes, which is important for small breeds.

84 Puerto Rican Slick-Haired Holstein Cattle Exhibit Enhanced Mammary Gland Hemodynamics Compared with Their Wild-Type Haired Counterparts

Abstract A mutation of the prolactin receptor gene (PRLR) results in cattle with short and sleek hair coat and superior thermoregulatory capacity when exposed to heat stress conditions. This mutation, which is present in Puerto Rican Holstein cattle (SLICK) results in decreased vaginal temperatures and increased milk yield (MY) compared with wild-type haired Holstein cattle (WT) under tropical conditions. The superior performance of SLICK Holstein cows has been considered multifactorial in nature and physiological differences between genotypes still need to be elucidated. Thereby, the objective was to compare the mammary gland hemodynamics and udder skin temperature between SLICK (n = 7) and WT (n = 7) Holstein cows during the summer as possible contributors to such physiological differences. Animals were selected for differences in MY (23.77±1.24 vs.19.74±1.24 kg, SLICK vs. WT, respectively; P = 0.041) and stratified by body weight (BW), lactation number, and days in milk (DIM). At 160±3 DIM, left and right pudendoepigastric arteries were assessed transrectally via Doppler ultrasonography and the udder skin temperature was determined using a thermal camera. Data were analyzed by one-way ANOVA using the MIXED procedure of SAS with resistance index (RI), pulsatility index (PI), diameter, total mammary blood flow (MBF), MBF relative to BW, and udder skin temperature as dependent variables and hair coat as the independent variable. The CORR procedure of SAS was used to determine the correlation between MY and MBF. Statistical significance was declared at P ≤ 0.05 and tendencies were established at 0.05 ≤ P ≤ 0.10. The RI was less (P = 0.045) in SLICK Holsteins (0.56 ± 0.02) than WT (0.64 ± 0.02). The PI tended (P = 0.0589) to be less in SLICK Holsteins (0.94±0.07) compared with WT (1.15 ± 0.07). The average diameter of the left and right pudendoepigastric arteries was greater in the SLICK animals compared with WT (1.31 ± 0.07 vs..08±0.07 cm, respectively; P = 0.038). Total MBF was increased in SLICK cattle compared with WT (9.03 ± 1.11 vs. 5.14 ± 1.11 L/min; P = 0.0291). There was a tendency (P = 0.0577) for the MBF relative to BW to be greater in the SLICK Holsteins compared with the WT (16.75 ± 2.14 vs.0.38 ± 2.14 mL/min*kg). No differences were observed in udder skin temperature between SLICK and WT Holsteins (P ≥ 0.05). Lastly, a tendency for a positive correlation between MY and MBF was observed (r = 0.51, P = 0.066). In the current study, SLICK Holsteins had enhanced blood supply to the mammary gland which is indicative of greater amount of oxygen and nutrients available for milk synthesis. Therefore, the gene introgression of the PRLR mutation to a dairy herd may result in animals with superior physiological performance during heat stress conditions preventing economical losses in the dairy industry.

Asymptomatic Familial Hyperprolactinemia Caused by a Unique bi-Allelic Variant in the Prolactin-Receptor Gene

Searchable abstracts of presentations at key conferences in endocrinology ISSN 1470-3947 (print) | ISSN 1479-6848 (online)

Alterations in gonadotropin, prolactin, androgen and estrogen receptor and steroidogenesis-associated gene expression in gander testes in relation to the annual period

Physiological mechanisms of seasonal changes in testicular function in birds are not fully elucidated. The balance between androgens and estrogens and testis sensitivity for gonadotropin and gonadal steroids are still unclear. The aim of the study was to examine: (1) the changes in circulating and intra-testicular steroid hormone levels and their relationship; (2) the mRNA expression of testicular gonadotropin, prolactin (PRL), progesterone (P4), androgen, and estrogen receptors, and (3) key steroidogenesis processes-related genes with immunofluorescent localization of aromatase in gander testes during the annual period. Testes from ganders (n = 25) in the first reproduction season were obtained at five breeding stages, i.e., prebreeding (PrB), peak of reproduction (PR), postbreeding (PoB), nonbreeding (NB), and onset of reproduction (OR). Males were kept under breeding conditions. It was found that plasma P4 levels decreased at the PoB and NB stages, whereas intra-testicular P4 was the highest in the NB stage. Intra-testicular estradiol (E2) levels were higher at the PoB and NB stages than the other stages, whereas testosterone (T) levels showed a nearly opposite pattern. The plasma estradiol–to–testosterone ratios were higher at the PrB, PoB and NB stages compared to other stages. The transcript abundances for luteinizing hormone receptor (LHR), PRL receptor (PRLR), estrogen receptor alpha (ERα), and estrogen receptor beta (ERβ) also change in testicular tissue during the annual period. Moreover, StAR mRNA expression was upregulated at the PoB and NB stages, and CYP11A1 transcript level was the highest at the PoB stage. Stage-dependent changes in the CYP19A1 mRNA and aromatase protein levels with higher abundances of transcript at PoB and NB stages and protein at the NB stage were observed. Localization and immunofluorescent signal intensity for aromatase also differed in relation to the examined stages. It may be suggested that differential E2 levels, as well as aromatase expression and localization across annual stages are responsible for the seasonal activation/inactivation stages of testis spermatogenesis in domestic ganders. These data strongly suggest a role of aromatase in the control of gander steroidogenesis as changes in this enzyme level are associated with alternation in gonadal steroid hormones. In addition, joint action with others hormones, like PRL and LH, seems to be important in the final effect of seasonal reproduction potential.

The Role of PRLR Gene Polymorphisms in Milk Production in European Wild Rabbit (Oryctolagus cuniculus).

One of the problematic points of rabbit breeding is that the nutritional requirements of the kits are not fully satisfied by the does' milk production from the third week of lactation onwards. The prolactin receptor gene has a significant effect on reproductive processes, and its polymorphisms have been associated with milk production in several species (cattle, goats, sheep, and buffalo). The European wild rabbit (Oryctolagus cuniculus), has a more diverse genetic background compared to domesticated lines. In the course of our study, sequencing of the 1210 bp long segment of the PRLR gene promoter region was accomplished. We detected four point mutations (SNP1-407G > A, SNP2-496G > C, SNP3-926T> and SNP4-973A > C) and one microsatellite at position 574. In our population, the four SNPs were segregated into four genotypes: AACCCCCC, GGGGTTAA, AAGGTTAC, and GGGGTCAC. Our results show that the genotype in the homozygous form is associated with higher milk production (1564.7 ± 444.7 g) compared to the other three genotypes (AACCCCCC 1399.1 ± 326.8 g; GTGACCTT 1403.8 ± 517.1 g; GGGGTCAC 1220.0 ± 666.2 g), and the short microsatellite repeat (167 bp) also coincides with significantly higher milk production (1623.8 ± 525.1 g). These results make the marker-assisted selection possible also for domesticated lines.

Study of population-genomic structure of Vyatka horses in interline aspect

Abstract. The population-genomic structure of the Vyatka horses was studied in the context of 10 male genealogical lines bred in the breed. Purpose. Evaluation of the genetic and breeding potential of the Vyatka breed, as well as the influence of factory breeds. Methods. Isolation of DNA from the hair follicles of Vyatka horses (n = 38) was performed using “ExtraGene DNA Prep 2000” by allele-specific PCR. Statistical calculations of allele frequencies and the types of studied loci were performed using MS Excel 10 software. Results. The polymorphism of MSTN, GYS1, DMTR3, CAST, and PRLR genes in Vyatka breed lines was studied for the first time. According to the frequency of occurrence CAST G/A (0.472) and PRLR G/C (0.417), as well as the genotype MSTN T/T (0.579) is somewhat dominated by the heterozygous genotypes characteristic of local breeds. They revealed polysaccharide accumulation mutation PSSM1 (0.189), characteristic of draft horses, and the DMRT3 mutation (0.087), which determines the ability for a non-standard gait - amble, noted among Oryol trotters, which indicates the presence of draft and trotting blood in the lines carrying these alleles. MSTN/C and DMRT3/A alleles, which are not typical for aborigines, are noted in Znatok line, whose representatives are distinguished by a lightweight body type and productive movements. The carriers of the mutant defective GYS1 allele (PSSM1) were identified in five lines: Bob, Gabizon, Buran, Dobrik, and Malakhit. Genotyping of all used stallions for genes associated with economically useful traits will allow more efficient selection in the Vyatka breed and prevent the spread of unwanted alleles, which is especially important for small breeds. Scientific novelty. For the first time, an intrapopulation genomic analysis of the domestic horse breed was studied in the interline aspect, and the influence of factory breeds on certain genealogical lines was shown. The polymorphism of the calpastatin (CAST) and prolactin receptor (PRLR) genes has not previously been studied in horse breeding.

Expression Variance of the PRLR Gene and its Association with Milk Productive Traits in Pure Holstein-Friesian and Montbeliarde×Holstein Crossbred Cattle Raised under Egyptian Condition

The objective of this study was to investigate the association between the expression level of the PRLR gene and milk production traits at different parities in pure Holstein-Friesian and Montbeliarde×Holstein crossbred cattle reared under Egyptian conditions, as well as its potential use in future breeding programs. For this purpose, eighteen healthy cows in mid lactation stage belonging to the first, second, and third parities were selected from each breed for sampling, RNA extraction, and quantitative real-time PCR. The results revealed that, the mean relative expression of prolactin receptor gene and its fold change level were significantly higher in pure Holstein-Friesian than Montbeliarde×Holstein crossbred at the level of the first three parities. In addition, there is a significant and strong positive correlation between PRLR1 and PRLR3 in pure Holstein-Friesian cattle. Significant negative correlations between PRLR2 and PRLR3, DMY3 and TMY3, were observed in Montbeliarde×Holstein crossbred cows. In conclusion, this information about the PRLR gene could be a useful indicator for the milk production trait and could be used in crossing programs that depend on genetic selection.

Mechanism underlying the modulation of milk production by incomplete milking.

Mammary gland secretory activity is modulated by systemic and local factors; however, the relationship between these factors is unknown. The aim of this study was to determine how a local factor, such as incomplete milking, affects mammary epithelial cell activity, number, and responsiveness to blood prolactin (PRL). Eight cows in mid-lactation were differentially milked (i.e., their right quarters were milked incompletely at approximately 70%, and their left quarters were milked completely, twice daily for 4 wk). Throughout the experiment, milk yield was measured at the quarter level. Milk samples were collected from each quarter once a week to assess the milk components, and epithelial cell concentrations, as well as to isolate milk fat globule RNA. In the weeks before and after the experiment, mammary gland functional capacity was evaluated by measuring the volume of milk harvested after complete filling of the gland. At the end of the last experimental week, mammary gland biopsies were performed on each rear quarter. The milk production of quarters milked completely remained stable during the treatment period, whereas, as expected, the milk production of quarters milked incompletely was only 53% of completely milked quarters at the end of the period. Accordingly, the expression of genes related to milk synthesis (CSN2, LALBA, and ACACA) in milk fat was lower in the quarters that were milked incompletely. Incomplete milking decreased the milk lactose content, indicating a loss of integrity of tight junctions. The total yield of epithelial cells in milk was not affected, but their concentration in milk, the BAX:BCL2 gene expression ratio, and the loss of mammary functional capacity were greater in the quarters milked incompletely, suggesting an acceleration of involution in those quarters. The expression of the short isoform of the PRL receptor gene (PRLR) tended to be lower, and the expression of STAT5A and STAT5B tended to decline in the quarters milked incompletely. In mammary gland biopsy samples, the number of both short and long isoforms of the PRLR were not affected, nor were the amount and activation of STAT3 and STAT5. However, the ratio of PRLR short isoform to PRLR long isoform was lower in the quarters milked incompletely. The decrease in milk yield induced by incomplete milking is rapid and associated with a decrease in mammary epithelial cell activity and a decrease in the number of secretory epithelial cells. The results of this experiment provide only limited support for the hypothesis that modulation of the mammary gland's responsiveness to PRL is part of the mechanism by which local factors, such as incomplete milking, modulate milk synthesis.