Tissue expression, association analysis between three novel SNPs of the RXRα gene and growth traits in Chinese indigenous cattle

Retinoid X receptor(RXR) α is a member of the nuclear hormone receptor superfamily that mediates the biological effects on several hormones,vitamins,and regulates lipid,glucose and energy metabolism.In this study,the tissue expression profiles of the bovine RXRαgene and association analysis of single nucleotide polymorphisms(SNPs) with growth traits were carried out in 413 Chinese native cattle.The expression profile was analysed in ten Jiaxian cattle tissues by real-time PCR,and the results showed that RXRαgene was abundantly expressed in adipose tissue and spleen,moderately expressed in heart,liver,lung,kidney,muscle and testis.Meanwhile,three SNPs(T27919A,T28139C and G28142A) and five haplotypes were identified.Haplotype with TTG was dominant with frequency of 69.1%.Chi-square test showed all populations were in Hardy-Weinberg equilibrium(P>0.05) at the three sites except Jiaxian cattle at G28142A site and Qinchuan cattle at T27919A site.Statistical analysis of combined sites showed that the individuals with TTGA genotype had significantly higher heart girth than those with TAGG genotype(P<0.05) and the animals with AAGA genotype had higher body weight than those with TAGG genotype(P<0.05) in T27919A-G28142A site.Heart girth,abdominal circumference and body weight of individuals with TCAG genotype were exceedingly higher than those with TTGG(P<0.01),TTGA and TCGG(P<0.05) in T28139C-G28142A site.For T27919A-T28139C site,the individuals of TCTA and TCTT genotype had significantly higher heart girth and lower height at hip cross than those with TTTA(P<0.05),and the body weight of TCAA and TCTT genotype individuals was higher than those with TTTA(P<0.05).In conclusion,these results provided evidence that the polymorphisms of RXRαgene were associated with growth traits and might apply to Chinese indigenous yellow cattle breeding program as a possible candidate for marker-assisted selection(MAS).     

Selection pressure drives the co-evolution of several lipid metabolism genes in mammals

Adipose tissue is an important endocrine organ and energy supplier.Its physiological effect on the regulation of the energy balance is considered an important factor underlying the evolution of mammals.To test whether the genes controlling lipid metabolism have undergone adaptive molecular change in the evolution of mammals,in this study,we used the orthologous gene sequences of 12 important lipid metabolism proteins (leptin,OB-RL,RXRA,RXRB,RXRG,PPARA,PPARB/D,PPARG,PNLIP,ADIPOQ,LPL and UCP1) from NCBI’s databases.We found evidence that 4 of the corresponding genes (leptin,ADIPOQ,PNLIP and PPARA) have undergone positive selection in their evolutionary history and that most adaptive changes occurred during the evolution of the super-clades Laurasiatheria (placentals) and suborders within Euarchontoglires (primates and rodents).Comparisons across sets of genes showed that in a third of cases,bursts of positive selection,more than would be expected by chance,occurred on corresponding branches.We propose that the positive selection drives adaptive changes in some lipid metabolism genes in or within Laurasiatheria and Euarchontoglires clades.Along with evidence from earlier studies,our results show that co-evolution among interacting lipid metabolism proteins has taken place.     

GFPT2基因系统发育与功能生物信息学分析

GFPT2作为己糖胺代谢重要的转录因子,在动物的多种疾病发生发展中起着重要作用。利用生物信息学方法对GFPT2蛋白的组分、功能、正选择位点及其分子进化进行了详细的分析。结果表明:13条不同物种氨基酸序列组分分析显示亮氨酸占比最高为9.86%,含量最低的氨基酸为色氨酸(0.15%),平均长度为664.8。系统进化树和结构域显示人和猴子的亲缘性最近,其次是大鼠、小鼠,所有氨基酸均包含两个SIS保守结构域。混合效应-演化模型(mixed effects model of evolution, MEME)和固定效应(fixed effects likelihood, REL)方法共发现8个正选择位点。基因本体论(gene ontology, GO)功能富集分析发现GFPT2主要参与GFPT酶活性、蛋白结合、碳水化合物衍生物结合方面发挥作用等生物学过程。京都基因与基因组百科全书(Kyoto encyclopedia of gene and genomes, KEGG)分析显示差异基因主要参与氨基糖和核苷酸糖代谢,丙氨酸、天冬氨酸和谷氨酸代谢,胰岛素抵抗3条信号通路。并且筛选到与GFPT2相互作用的10个基因:GFPT1、GLUL、GNPDA1、GNPNAT1、GPI、HK1、MPI、PPAT、AMDHD2、CAD。研究结果可以为GFPT2分子功能的深层次研究提供一定的借鉴作用,对进一步探究由己糖胺代谢导致的代谢异常及心血管疾病治疗具有重要的现实意义。     

The effect of pmpR on the type III secretion system in Pseudomonas aeruginosa

The type III secretion system(T3SS) plays important roles in Pseudomonas aeruginosa pathogenicity.Previously,we reported that the uncharacterized protein PmpR could regulate pqsR,an important regulator in the quorum-sensing system,by directly binding to its promoter region.As the T3SS is controlled by the quorum-sensing system,here,we investigated the relationship between PmpR and the T3SS.Our data showed that expression of the T3SS genes exoS,exoY,exoT,and exsD was dramatically increased in a pmpR-deletion mutant compared with that in the wild-type P.aeruginosa strain PAO1.Data from DNA mobility assays indicated that PmpR affects the T3SS indirectly.It is unlikely that PmpR controls the T3SS via the Pseudomonas quinolone signal(PQS) because the PQS negatively regulates the T3SS,while pmpR negatively regulates the PQS.The effect of PmpR on the T3SS seems to be independent of the PQS;further investigation is required to uncover the underlying regulatory pathways.     

Fluorescent co-localization of PTS1 and PTS2 and its application in analysis of the gene function and the peroxisomal dynamic in Magnaporthe oryzae

The peroxisomal matrix proteins involved in many important biological metabolism pathways in eukaryotic cells are encoded by nucleal genes, synthesized in the cytoplasm and then transported into the organelles. Targeting and import of these proteins depend on their two peroxisomal targeting signals （PTS 1 and PTS2） in sequence as we have known so far. The vectors of the fluorescent fusions with PTS, i.e., green fluorescence protein （GFP）-PTS1, GFP-PTS2 and red fluorescence protein （RFP）-PTS1, were constructed and introduced into Magnaporthe oryzae Guy ll cells. Transformants containing these fusions emitted fluorescence in a punctate pattern, and the locations of the red and green fluorescence overlapped exactly in RFP-PTS 1 and GFP-PTS2 co-transformed strains. These data indicated that both PTS1 and PTS2 fusions were imported into peroxisomes. A probable higher efficiency of PTS1 machinery was revealed by comparing the fluorescence backgrotmds in GFP-PTS1 and GFP-PTS2 transformants. By introducing both RFP-PTS1 and GFP-PTS2 into Amgpex6 mutants, the involvement of MGPEX6 gene in both PTS1 and PTS2 pathways was proved. In addition, using these transformants, the inducement ofperoxisomes and the dynamic of peroxisomal number during the pre-penetration processes were investigated as well. In summary, by the localization and co-localization of PTS1 and PTS2, we provided a useful tool to evaluate the biological roles of the peroxisomes and the related genes.     

Metabolism of fibrates by cytochrome P450s and UDP-glycosyltransferases in rat and human liver microsomes

Fibrates are widely used for the treatment of dyslipidemia.However,the contributions of the phase I and phase II metabolic pathways to the clearance of fibrates are unclear.In this study,we investigated the metabolism of gemfibrozil(Gem) ,clofibric acid(CA) ,fenofibric acid(FA) and bezafibrate(Beza) by cytochrome P450s(P450s) and UDP-glycosyltransferases(UGTs) using a substrate depletion approach.We also compared the metabolic characteristics of rat liver microsomes(RLM) and human liver microsomes(HLM) .The intrinsic clearance rates mediated by P450s,UGTs and both were 172 ± 22,643 ± 26,798 ± 103 μL min-1 mg-1,respectively,for Gem and 43 ± 11,88 ± 12,119 ± 15 μL min-1 mg-1,respectively,for CA in RLM.The fractions metabolized by P450s and UGTs in RLM were 22% and 81% for Gem,36% and 74% for CA.The P450-and UGT-mediated depletion rates for Gem were 303 and 1607 nmol min-1 mg-1 in RLM versus 86 and 243 nmol min-1 mg-1 in HLM.The corre-sponding rates for CA were 1.1 and 1.7 nmol min-1 mg-1 in RLM versus 0.025 and 0.038 nmol min-1 mg-1 in HLM.Accordingly,both P450s and UGTs substantially contribute to the clearance of Gem and CA,with UGTs playing a greater role.To avoid un-der-estimating the impact of these pathways,it is necessary to measure NADPH-and UDPGA-dependent metabolism.Although the fractions of these two pathways in RLM and HLM were similar,the depletion rate of Gem and CA in RLM was higher than that in HLM.The metabolism of FA and Beza by P450s and UGTs was too low to calculate intrinsic clearance in both RLM and HLM.These results indicate that fibrates are metabolized via similar pathways in rats and humans,and it is applicable to use RLM to predict the clearance of fibrates in human.     

JAK2在生长激素介导的信号传导中的作用

在细胞水平上，JAK2在生长激素介导的信号传导中具重要作用。生长激素与生长激素膜蛋白受体结合，激活胞质酪氨酸激酶JAK2后，JAK2自身磷酸化。同时磷酸化生长激素膜蛋白受体，从而形成信号传导因子与转录激活因子、适配蛋白Shc等细胞信号分子高亲和位点。生长激素刺激下的JAK2也会磷酸化胰岛素受体底物，从而激活磷酯酰肌糖3激酶以及其它相关的调节新陈代谢的生物分子活性。而且JAK2还能激活适配蛋白SH2-B。这些因子和激活途径可能是生长激素作用于机体并调节机体生长代谢的基础。     

全基因组关联分析显示基因ANXA8和C10orf11为影响肌少症的候选基因

为了寻找与瘦体重(lean body mass,LBM)相关的单核苷酸多态性(single nucleotide polymorphism, SNP)位点及易感基因,在1 000个不相关的白人中采用Affymetix 500K芯片扫描了500 000个SNPs,并进行全基因组关联分析(genome-wide association study,GWAS),显著结果在1 625个中国人样本和2 283个欧洲白人样本中进行验证,并将验证结果与研究结果进行荟萃分析。研究发现SNPsrs7905603,rs9416083,rs4409772,rs2894310与LBM关联,其中rs7905603位于基因ANXA8,其他3个SNPs位于基因C10orf11。荟萃分析得到的合并p值分别为2.08×10-5,7.44×10~(-6),6.73×10~(-6),6.76×10~(-6)。ANXA8和C10orf11基因是影响LBM变异的候选基因,这对肌少症的认识提供了新的理论依据。     

菠菜转录因子MYB基因家族的鉴定及表达分析

利用生物信息学手段鉴定了76个具有典型R结构的菠菜转录因子（Spinacia oleracea） MYB,其中包括72条R2R3-MYB基因（2R-MYB）和4条R1R2R3-MYB基因（3R-MYB）。通过生物信息学对菠菜MYB转录因子家族成员的理化性质、染色体定位、结构域序列保守性和系统进化关系进行了分析,结果表明：菠菜MYB家族有32个基因位于染色体正链,另外44个基因位于染色体反链;MYB的DNA结合域中的保守域主要位于两个R重复序列的第二和第三螺旋之间,结合域中每个R重复的第一和第二色氨酸之间的氨基酸序列相对不保守;根据菠菜、拟南芥及甜菜的MYB家族系统进化关系可以推测,菠菜MYB家族中56个成员可以按功能划分为4类,在菠菜的生长发育过程中可能起着重要的调节作用,其余成员中有7个MYB基因可能参与菠菜响应氮素浓度的氮素利用及生长发育进程。     

基于网络药理学的蒲黄对血瘀证的作用靶点与代谢通路研究

本研究利用网络药理学探讨蒲黄（Pollen typhae）治疗血瘀证的作用靶点与代谢通路。研究首先通过PharmGKB、TTD和CTD数据库筛选出与血瘀证相关的靶点;接着利用String平台构建蛋白质相互作用（PPI）网络,并挖掘PPI网络中潜在的蛋白质功能模块,通过Metascape平台分析蒲黄活性成分作用靶点所参与的生物过程及通路;然后采用Cytoscape3.7.2软件构建成分-靶点网络、疾病-靶点网络、成分-靶点-通路网络;最后通过AutoDock Vina软件将蒲黄活性成分、阳性药阿司匹林（Aspirin）与核心靶点进行分子对接验证,并比较各自之间的对接强度。结果表明,槲皮素、山奈酚、异鼠李素、异鼠李素-3-O-新橙皮苷等在参与的成分-靶点网络、疾病-靶点网络、成分-靶点-通路网络中联系密切,其与人体相互作用强,可能是治疗血瘀证的活性成分;核心作用靶点有血管内皮生长因子（VEGFA）、蛋白激酶（AKT1）、雌激素受体（ESR1）、肿瘤坏死因子（TNF）、转录因子（JUN）、有丝分裂原激活蛋白激酶14（MAPK14）等。蒲黄治疗血瘀证的生物学通路主要涉及VEGF信号通路（VEGF signaling pathway）,IL-17信号通路（IL-17 signaling pathway）,NF-κB信号通路（NF-kappa B signaling pathway）等,其治疗方面主要涉及血管内皮生长因子、血管功能和血液循环等。分子对接验证显示,活性成分和作用靶点的结合能小于-5的占85.88％,即大部分靶点与槲皮素、山奈酚、异鼠李素、异鼠李素-3-O-新橙皮苷的结合活性较好;而Aspirin与靶点的结合强度较蒲黄活性成分异鼠李素-3-O-新橙皮苷、（2R）-5,7-二羟基-2-（4-羟基苯基）苯并吡喃-4-酮、儿茶素和表儿茶素的差。通过网络药理学初步揭示蒲黄多成分、多靶点、多通路的作用特点,预测了蒲黄治疗血瘀证的可能作用靶点和代谢通路;且从分子对接结果来看,蒲黄活性成分异鼠李素-3-O-新橙皮苷、（2R）-5,7-二羟基-2-（4-羟基苯基）苯并吡喃-4-酮、儿茶素和表儿茶素等与核心靶点的对接效果均优于阳性药Aspirin。