Differential promoter activities of functional haplotypes in the 5′‐flanking region of human Sulfotransferase 1A1

Previously, we reported five common single nucleotide polymorphisms (SNPs), ?624G>C, ?396G>A, ?358A>C, ?341C>G, and ?294T>C, and six common haplotypes (CGACT, GAACT, GGAGC, GGACC, CAACT, and GAACC) in the 5′‐flanking region of the SULT1A1 gene that were associated with altered enzymatic activity. In the present study, we performed in vitro assays to determine the functional impact of these genetic variations on the promoter activity. Dual luciferase reporter assays revealed that these SNPs are located in a negative regulatory fragment of the SULT1A1 gene. Further experiments demonstrated that these SNPs and haplotypes affected promoter activities of SULT1A1. Electrophoretic mobility shift assays showed distinctive binding patterns for the SNPs ‐396G>A and ‐294T>C, due to differential binding affinities of the G/A alleles and the T/C alleles to nuclear proteins extracted from the liver carcinoma cell lines, HepG2 and Huh7. © 2012 Wiley Periodicals, Inc. J Biochem Mol Toxicol 26:422–428, 2012; View this article online at wileyonlinelibrary.com . DOI 10:1002/jbt.21437     

SARS冠状病毒S蛋白候选细胞受体氨基肽酶N的基因变异研究

氨基肽酶N是人类冠状病毒HCoV 2 2 9E的细胞受体 ,可能为SARS冠状病毒 (SARS CoV)S蛋白的受体。应用变性高效液相色谱 (DHPLC)技术筛查了正常无关个体中氨基肽酶N编码基因ANPEP的全部外显子及其两侧部分内含子序列。对筛查中DHPLC峰型提示有基因变异存在的DNA片段行PCR产物直接测序 ,共发现 9种单核苷酸多态 (SNP) ,其中 4种为非同义SNP ,分别为T32 1M(96 2C >T)、S6 5 1L(195 2C >T)、S75 2N(2 2 5 5G >A)和G76 4R(2 2 90G >A) ;其余 5种SNP为T795T(2 385C >T)、IVS7+17G >A、IVS14 16A >G、IVS17+12C >G和IVS17+44C >T。这些SNP的发现 ,为SARS CoV的宿主遗传因素研究 ,特别是发现SARS CoV感染和SARS发病的易感基因或抗病基因 ,提供了遗传标记。     

The association of SNPs in Hsp90β gene 5′ flanking region with thermo tolerance traits and tissue mRNA expression in two chicken breeds

Thermo stress induces heat shock proteins (HSPs) expression and HSP90 family is one of them that has been reported to involve in cellular protection against heat stress. But whether there is any association of genetic variation in the Hsp90β gene in chicken with thermo tolerance is still unknown. Direct sequencing was used to detect possible SNPs in Hsp90β gene 5′ flanking region in 3 chicken breeds (n = 663). Six mutations, among which 2 SNPs were chosen and genotypes were analyzed with PCR–RFLP method, were found in Hsp90β gene in these 3 chicken breeds. Association analysis indicated that SNP of C.?141G>A in the 5′ flanking region of the Hsp90β gene in chicken had some effect on thermo tolerance traits, which may be a potential molecular marker of thermo tolerance, and the genotype GG was the thermo tolerance genotype. Hsp90β gene mRNA expression in different tissues detected by quantitative real-time PCR assay were demonstrated to be tissue dependent, implying that different tissues have distinct sensibilities to thermo stress. Besides, it was shown time specific and varieties differences. The expression of Hsp90β mRNA in Lingshan chickens in some tissues including heart, liver, brain and spleen were significantly higher or lower than that of White Recessive Rock (WRR). In this study, we presume that these mutations could be used in marker assisted selection for anti-heat stress chickens in our breeding program, and WRR were vulnerable to tropical thermo stress whereas Lingshan chickens were well adapted.     

Single-nucleotide polymorphisms in the promoter of the growth hormone-releasing hormone receptor gene are associated with growth and reproduction traits in chickens

Growth hormone‐releasing hormone receptor (GHRHR) plays a critical role in growth hormone (GH) synthesis, release and regulation in animals. The objective of this study was to investigate variations of the chicken GHRHR gene and their associations with growth and reproduction traits in 768 Beijing You chickens. Results revealed three single nucleotide polymorphisms (SNPs) in the promoter region of the gene (g.‐1654A>G, g.‐1411A>G and g.‐142T>C). Association analysis revealed that the novel SNP g.‐1654A>G had significant effects on chicken body weight at 7, 9, 11, 13, 17 weeks of age and the age of first egg as well as egg number at 32, 36 and 40 weeks. Significant association was also observed between g.‐1411A>G and g.‐142T>C with EN24. Moreover, the age of first egg was distinctly related with g.‐142T>C (P < 0.05). Although significant statistical difference was not detected in GHRHR mRNA levels among genotypes of the SNPs (P > 0.05), strong expression variations of the gene were found between the ages 17 and 20 weeks in the population (P < 0.05). These results suggest that the three SNPs in the GHRHR promoter could be used as potential genetic markers to improve the growth and reproductive traits in chickens.     

延边朝鲜族和汉族脂联素启动子SNPs与原发性高血压的相关性

为了探讨延边朝鲜族和汉族脂联素基因启动子单核苷酸多态性(SNPs)与原发性高血压(EH)的关系, 文章采用PCR产物直接测序方法检测了220例EH患者和268例对照个体的脂联素启动子5个SNPs位点: -11426A>G(rs16861194)、-11391G>A(rs17300539)、-11377C>G(rs62620185)、-11156insCA(rs60806105)、-11043C>T(rs76786086), 氧化酶法测定空腹血糖、甘油三酯、总胆固醇、低密度脂蛋白、高密度脂蛋白, 酶联免疫吸附法(ELISA)测定血浆脂联素和胰岛素。结果显示: (1) -11426A>G、-11377C>G 和-11156insCA 3个位点具有多态性, 且它们的基因型频率分布符合Hardy-Weinberg平衡定律(P>0.05), -11391G>A和-11043C>T位点无多态性; (2) -11426A>G和-11156insCA呈完全连锁不平衡(D’=1; r2=1); (3) -11426G基因频率比较, 朝鲜族(21.10%)高于汉族(12.05%), 汉族EH组高于对照组; -11377C>G的基因型和基因频率在朝鲜族和汉族间及同一民族内EH组和对照组间比较均无统计学意义(P>0.05); (4)单倍型?11426G -11377C的频率, 汉族EH组高于对照组(P<0.05), 朝鲜族EH组和对照组比较无统计学意义(P>0.05); (5)EH组的血浆脂联素水平明显低于对照组(P<0.001)。据此得出结论: (1)首次发现?11426A>G和?11156insCA呈完全连锁不平衡, -11426 A>G的多态性在朝鲜族和汉族中存在民族差异; (2) -11426 G和-11426G -11377C是延边汉族EH的危险因子和危险单倍型, 但不是朝鲜族的; (3)低血浆脂联素是延边朝鲜族和汉族EH的重要危险因素; (4)血浆脂联素水平与-11426A>G基因型无关。     

Polymorphisms of the TIM-1 gene are associated with rheumatoid arthritis in the Chinese Hui minority ethnic population

The T-cell immunoglobulin and mucin domain 1 (TIM-1) is known to be associated with susceptibility to rheumatoid arthritis (RA). We investigated the association of four single-nucleotide polymorphisms (SNPs) in the promoter region of the TIM-1 gene with susceptibility to RA in a Chinese Hui ethnic minority group. Using RFLP or sequence specific primer-PCR, 118 RA patients and 118 non-arthritis control individuals were analyzed for the -1637A>G, -1454G>A, -416G>C, and -232A>G SNPs in the TIM-1 gene. The polymorphisms -232A>G and -1637A>G in the promoter region of TIM-1 were found to be associated with susceptibility to the RA gene in the Hui population, while -416G>C and -1454G>A SNPs were not. Of these, the polymorphism of -232A>G is inconsistent with that found in a Korean population, suggesting that genetic variations of the TIM-1 gene contribute to RA susceptibility in different ways among different populations. Based on haplotype analysis, individuals with haplotypes AGCA (Χ(2) = 22.0, P < 0.01, OR (95%CI) >1), AGCG (Χ(2) = 18.16, P < 0.01, OR (95%CI) >1) and AGGA (Χ(2) = 5.58, P < 0.05, OR (95%CI) >1) are at risk to develop RA in the Chinese Hui population; those with the GAGA (Χ(2) = 7.44, P < 0.01, OR (95%CI) <1) haplotype may have a decreased likelihood of RA. GGCA and GGCG are more common in both RA and non-RA subjects. We conclude that -1637A>G and -232A>G polymorphisms of TIM-1 are associated with susceptibility to RA in the Chinese Hui population.     

A new single nucleotide polymorphism in the rabbit (Oryctolagus cuniculus) myostatin (MSTN) gene is associated with carcass composition traits

This study aimed at the identification of genetic variations in the myostatin (MSTN) gene and testing their effects on carcass quality traits. We comparatively sequenced Giant Grey (GG) and New Zealand White (NZW) rabbits that were founders of a cross‐bred population. Alignment of our sequence data with the GenBank sequence of the rabbit MSTN gene (Ensembl Gene ID ENSOCUG00000012663) identified three single nucleotide polymorphisms (SNPs). The two novel SNPs (c.?125T>C, c.373+234G>A) and one known SNP (c.747+34C>T) were subsequently analysed for linkage with carcass composition traits in 363 F₂ animals of the cross GG × NZW. Significant linkage was found between c.373+234G>A and nine carcass composition traits (P < 0.05). No significant effects were found for c.?125T>C and c.747+34C>T. Because the linked SNP is located in intron 1 and no genetic variation was found in the coding region, further investigations are necessary to understand the functional effect of the c.373+234G>A variant on the variability of the traits.     

Three novel SNPs of the bovine Tf gene in Chinese native cattle and their associations with milk production traits

Transferrin (Tf) is a β-globulin protein that transports iron ions in mammalian cells. It contributes to innate immunity to microbial pathogens, primarily by limiting microbial access to iron. Thus, polymorphisms present in bovine Tf could potentially underlie inherited differences in mastitis resistance and milk production traits. We detected three novel single-nucleotide polymorphisms of the Tf gene in Chinese native cattle by screening for genetic variation of Tf in 751 individuals of three Chinese cattle breeds, namely China Holstein, Luxi Yellow and Bohai Black, using PCR-RFLP and DNA sequencing techniques. The three new SNPs, g.-1748G>A ss250608649, g.13942T>C ss250608650, and g.14037A>G ss250608651, had allele frequencies of 85.9, 86.3 and 92.5%, 64.5, 73.3 and 65.0%, and 67.6, 73.7 and 60.0%, respectively. SNP g.-1748G>A was located in the 5' flanking region of Tf. SNP g.14037A>G was located in intron 8 of Tf. SNP g.13942T>C, located in exon 8 of Tf, was a synonymous mutation (TTA > CTA), encoding a leucine (326 aa) in the Tf protein. Associations of the Tf SNPs with milk traits were also analyzed. Significant (P < 0.05) relationships among the Tf polymorphisms, somatic cell scores (SCS), and milk productive traits were observed. Cows with genotypes TT (g.13942T>C), GG (g.-1748G>A) and AG (g.14037A>G) had a lower SCS and higher protein levels and 305-day milk yield. Nineteen combinations of different haplotypes from the three SNPs were identified in Chinese Holstein cattle. The haplotype combination ATA/GCA, GCA/GCA and GCG/ GTA was dominant in cows with a lower SCS, a higher protein level and a higher 305-day milk yield, respectively. Moreover, the gene expression level of Tf was higher in mastitis-affected mammary tissues than in normal mammary tissues. These results suggest that the Tf gene affects milk production, as well as mastitis-resistance traits, in Chinese Holsteins.     

Detection of Bovine Central Nervous System Tissue as Bovine Spongiform Encephalopathy Risk Material by Real-Time Reverse Transcriptase-PCR in Raw and Cooked Beef Products

PCR-SSCP and DNA sequencing methods were applied to reveal single nucleotide polymorphisms (SNPs) in the bovine VEGF-B gene in 675 samples belonging to three native Chinese cattle breeds. We found 3 SNPs and a duplication NC_007330.5: g. [782 A>G p. (Gly112 =) (;) 1000-1001dup CT (;) 1079 C>T (;) 2129 G>A p. (Arg184Gln)]. We also observed a statistically significant association of the polymorphism (1000-1001dup CT) in intron 3 of the VEGF-B gene with the body weight of the Nanyang cattle (p < 0.05). This polymorphisms of VEGF-B gene need to be verified among a larger cattle population before it can be identified as a marker for bovine body weight.     

Novel SNPs of the Bovine LEPR Gene and Their Association with Growth Traits

In this study, polymorphism in the bovine LEPR gene exon 4 was detected by PCR-SSCP and DNA sequencing methods in 653 individuals from five Chinese cattle breeds. Two haplotypes (M and N), three observed genotypes (MM, MN, and NN), and five single nucleotide polymorphisms (SNPs) (NC_007301:g.26767T>C, NC_007301:g.26805C>T, NC_007301:g.27050A>G, NC_007301:g.27063G>A, NC_007301:g.27079G>A) were detected. The frequencies of haplotypes M and N in the five breeds were 0.661-0.747 and 0.253-0.339, respectively. The SNP locus was in Hardy-Weinberg equilibrium in Nanyang, Jiaxian red, Angus, and Jinnan cattle (P > 0.05) and was in Hardy-Weinberg disequilibrium in Qinchuan cattle (P < 0.05). Polymorphism of the LEPR gene was shown to be associated with growth traits in the Nanyang breed. The SNP in the bovine LEPR gene had significant effects on body height, body length, body weight, heart girth, and average daily gain at 6 and 12 months old (P < 0.01 or P < 0.05). Therefore, these results suggest that the LEPR gene is a strong candidate gene that affects growth traits in cattle.     

Polymorphisms of the Myostatin Gene and Its Relationship with Reproduction Traits in the Bian Chicken

Myostatin, or growth and differentiation factor 8, is a member of the transforming growth factor-β superfamily; it functions as a negative regulator of skeletal muscle development and growth in mammals. In this study, single nucleotide polymorphisms in the 5′ regulatory region and exon 1 of the myostatin gene were detected by PCR–SSCP in the Bian, Jinghai, Youxi, and Arbor Acre chickens, and the associations of the polymorphisms with reproduction traits were analyzed. Seven SNPs (A326G, C334G, C1346T, G1375A, A1473G, G1491A, and G2283A) were found in the myostatin gene. Association analysis showed that the G2283A were significantly associated with reproduction traits. Bian chickens of the GG genotype had a greater age at first egg than those of the GA and AA genotypes (P < 0.01). Correspondingly, Bian chickens of the GA and AA genotypes had larger egg number at 300 days than those of the GG genotype (P < 0.05 and P < 0.01, respectively). Bian chickens of the AA genotype had significantly higher body weight at 300 days than those of the GG genotype (P < 0.05). These results suggested that the myostatin gene may have certain effects on reproduction traits other than merely as a negative regulator of skeletal muscle development and growth in mammals previously reported.     

Genome-Wide Association Study Identifies Loci and Candidate Genes for Body Composition and Meat Quality Traits in Beijing-You Chickens

Body composition and meat quality traits are important economic traits of chickens. The development of high-throughput genotyping platforms and relevant statistical methods have enabled genome-wide association studies in chickens. In order to identify molecular markers and candidate genes associated with body composition and meat quality traits, genome-wide association studies were conducted using the Illumina 60 K SNP Beadchip to genotype 724 Beijing-You chickens. For each bird, a total of 16 traits were measured, including carcass weight (CW), eviscerated weight (EW), dressing percentage, breast muscle weight (BrW) and percentage (BrP), thigh muscle weight and percentage, abdominal fat weight and percentage, dry matter and intramuscular fat contents of breast and thigh muscle, ultimate pH, and shear force of the pectoralis major muscle at 100 d of age. The SNPs that were significantly associated with the phenotypic traits were identified using both simple (GLM) and compressed mixed linear (MLM) models. For nine of ten body composition traits studied, SNPs showing genome wide significance (P<2.59E−6) have been identified. A consistent region on chicken (Gallus gallus) chromosome 4 (GGA4), including seven significant SNPs and four candidate genes (LCORL, LAP3, LDB2, TAPT1), were found to be associated with CW and EW. Another 0.65 Mb region on GGA3 for BrW and BrP was identified. After measuring the mRNA content in beast muscle for five genes located in this region, the changes in GJA1 expression were found to be consistent with that of breast muscle weight across development. It is highly possible that GJA1 is a functional gene for breast muscle development in chickens. For meat quality traits, several SNPs reaching suggestive association were identified and possible candidate genes with their functions were discussed.     

Resistin polymorphisms are associated with muscle, bone, and fat phenotypes in white men and women

Objective: The biological function of resistin (RST) is unknown, although it may have roles in obesity, diabetes, and insulin resistance. The objective of this study was to examine the effects of single nucleotide polymorphisms (SNPs) in the human RST gene on muscle, bone, and adipose tissue phenotypes and in response to resistance training (RT). Research Methods and Procedures: Subjects were white and consisted of strength (n = 482) and size (n = 409) cohorts who had not performed RT in the previous year. Subjects completed 12 weeks of structured, unilateral upper arm RT aimed at increasing the size and strength of the non‐dominant arm, using their dominant arm as an untrained control. Strength measurements were taken pre‐ and post‐12‐week RT and consisted of elbow flexor isometric strength and one‐repetition maximum during a biceps curl using free weights. Whole muscle, subcutaneous fat, and cortical bone volumes were measured by magnetic resonance imaging. Six RST SNPs were identified. Analysis of covariance was used to test for effects of the SNPs on pre‐ and post‐muscle strength and whole muscle, fat, and bone volumes independent of gender, age, and body weight. Results: Five RST SNPs (?537 A>C, ?420 C>G, 398 C>T, 540 G>A, 980 C>G) were associated with measured phenotypes among subjects when stratified by BMI (<25, ≥25 kg/m²). Several gender‐specific associations were observed between RST SNPs and phenotypes among individuals with a BMI ≥ 25. Conversely, only two associations were observed among individuals with a BMI < 25. Discussion: These data support previous identified associations of RST with adipose tissue and demonstrate additional associations with bone and skeletal muscle that warrant further investigation.