小鼠富亮氨酸重复结构蛋白Lrig2 基因、蛋白结构及组织定位分析

目的:分析小鼠富亮氨酸重复结构蛋白家族成员Lrig2的基因与蛋白的结构,明确其组织分布和定位,并对其功能进行初步预测。方法:利用生物信息学分析技术对小鼠Lrig2基因的染色体定位、蛋白结构进行分析预测;通过RT-PCR、mRNA原位杂交技术检测Lrig2基因在小鼠不同组织中的表达定位;通过系统进化树分析Lrig2与其他Lrrs蛋白家族成员的同源性。结果:生物信息学分析显示,Lrig2是一种跨膜蛋白受体,是Lrrs蛋白超家族成员之一,胞外区含有15个Lrr模序、3个免疫球蛋白样结构域,存在单一的跨膜结构域;Lrig2在小鼠的多个组织中表达,其中在胸腺、脾脏等组织中表达较强;系统树分析显示,Lrigs蛋白是sLrPs超家族成员,是一种跨膜蛋白受体。结论:Lrig2在免疫组织中表达较强,推测其可能在肿瘤免疫应答进程中发挥重要的效能。     

烟草C2H2锌指蛋白转录因子家族成员的鉴定与表达分析

C2H2锌指蛋白转录因子家族在真核生物中具有重要的生物学功能,广泛参与植物叶的发生、花器官的调控、侧枝的形成及逆境胁迫等生命过程。植物C2H2锌指蛋白不仅结合DNA和RNA,而且与蛋白质之间相互作用。本研究利用普通烟草(Nicotiana tabacum)基因组数据库,运用Blastp比对,结合Pfam和SMART分析,鉴定了118条普通烟草C2H2锌指蛋白家族成员;对烟草C2H2锌指蛋白家族进行了进化树分析、结构域分析、物理化学性质分析、染色体定位、基因结构分析、三维结构分析及组织表达分析等。结果表明：不同成员的氨基酸长度差异较大;系统进化及结构域分析显示,所有C2H2家族成员可以被分为5个亚家族,同一亚家族成员之间在结构域和理化性质上呈现较高一致性;每个成员都含有C2H2结构域,在数量上存在较大差异;将所有基因家族成员定位在22条染色体上;组织表达分析表明,每个C2H2亚家族都有成员在不同组织中表达,在叶及根中有些基因的表达量较高。     

小鼠脂联素受体2基因编码区的克隆及序列分析

构建小鼠脂联素受体2(mAdipoR2)基因cDNA克隆,并进行序列及基因结构分析,为进一步研究小鼠AdipoR2的表达和生物学活性奠定基础。用RT—PCR方法扩增mAdipoR2基因cDNA,获得的片段连接至pGEM-T载体,转化JM109大肠杆菌,经酶切鉴定后,进行序列测定。利用因特网生物信息学资源分析mAdipoR2基因序列。结果成功地构建了mAdipoR2基因cDNA克隆,其序列与GenBank登录序列一致。通过与小鼠基因组序列比对,分析了mAdipoR2基因结构,其编码序列由7个外显子组成,编码1个含7个跨膜区的膜蛋白,但该受体不属于G蛋白偶联受体家族(GPCRs)。mAdipoR2与人及大鼠蛋白的同源性分别为91％和95％。     

家鸡G蛋白偶联受体85基因的结构、序列与组织表达分析

G蛋白偶联受体(GPCR)超家族是细胞膜上广泛存在的一类受体,是细胞跨膜信号转导的一类重要受体分子,参与许多生理过程调节。它们中仍有很多至今尚未找到内源性配体,这类受体被称为孤儿型受体。G蛋白偶联受体85(GPR85)是GPCR超家族中孤儿型受体的一员。目前,在非哺乳类脊椎动物中,针对GPR85的研究极少。本研究以家鸡Gallus gallus domesticus为模型,通过反转录PCR和RACE-PCR等方法从脑中克隆到GPR85基因的cDNA全长序列,揭示其基因结构,并用实时荧光定量PCR(qPCR)方法探究了该基因在家鸡各组织中的表达情况。结果显示:家鸡GPR85基因位于1号染色体上,由2个外显子组成,其编码区位于第2个外显子上,长为1 113 bp,可编码1个370个氨基酸的7次跨膜受体蛋白。家鸡GPR85与其他脊椎动物(人Homo sapiens、小鼠Mus musculus、大鼠Rattus norvegicus、热带爪蟾Xenopus tropicalis和斑马鱼Danio rerio)的GPR85具有高度的氨基酸序列一致性(>93%)。qPCR分析发现,GPR85基因mRNA在家鸡全脑、垂体、肾上腺、精巢中有较高表达,而在所检测的其他外周组织中表达极低。本研究首次揭示了家鸡GPR85基因的结构与表达特征,为后续探究GPR85基因在家鸡等非哺乳类中的生理功能奠定基础。     

人p17.3基因及其推导蛋白的结构和功能分析

用生物信息学技术研究人类神经元蛋白p17.3的基因结构、染色体定位、组织表达特征及其推导蛋白的理化特性、空间结构和功能等。以高通量基因组序列(HTGS)数据库及SAGE文库为基础对p17.3基因进行染色体定位及组织表达谱分析。通过DAS和TMpred等程序预测其蛋白质结构及功能。利用PDB程序模拟其三维空间结构。研究表明p17.3基因定位于人类染色体Xp11.2-3,在多种组织中广谱表达,但在前列腺表达量最高。其编码蛋白含有一个PEST区域、一个线粒体靶向序列和一个潜在“跨膜螺旋”结构。p17.3基因可能是一个参与神经元发育调控的基因。     

人Transgelin蛋白家族生物信息学分析

本研究通过对人Transgelin蛋白家族(Transgelins)3位成员(Transgelin,-2,-3)的生物信息学预测分析,显示Transgelin和-2等电点分别为8.87和8.41,Transgelin-3等电点为6.84,3位成员均为非分泌型蛋白,且极有可能是跨膜型蛋白.蛋白质稳定性分析结果显示,除Tr...     

家鸡G蛋白偶联受体78基因的克隆与组织表达图谱分析

G蛋白偶联受体(GPCR)是一大类膜受体超家族,参与了机体几乎所有的生理过程,是一类重要信号分子受体。GPR78作为GPCR超家族的成员之一,属于孤儿型受体。目前,在脊椎动物中,针对该基因结构与功能的研究极少。本研究以家鸡为动物模型,通过RT-PCR方法克隆了GPR78基因的编码区序列,并探究了该基因在家鸡各组织中的表达情况。结果显示:家鸡GPR78基因编码区全长为1020 bp,含3个外显子,可编码1个长为339个氨基酸的7次跨膜受体;该基因在家鸡脑各功能区及垂体中高表达,而在其他外周组织中均不表达。本研究为后续探究GPR78基因在家鸡中的生理功能奠定基础。     

人P2X3蛋白结构与功能的生物信息学分析

嘌呤能受体P2X3(P2X3 receptor,P2X3R)为一种双跨膜蛋白,在疼痛、咳嗽等涉及神经信号传导的疾病中发挥作用.本研究利用生物信息学技术对人P2X3R进行理化性质、B细胞抗原表位、同源性、信号肽和跨膜区、二级结构、互作蛋白、磷酸化修饰以及基因本体论预测和分析,发现其主要表达于神经系统,常以P2X2/3R异...     

一条大鼠睾丸新基因的生物信息学分析及真核表达

目的：探讨大鼠睾丸组织一条新基因的生物信息学特征和真核表达。方法：构建pEGFP-N1载体的融合质粒进行真核表达,利用生物信息学手段分析基因和蛋白功能。结果：生物信息学分析表明RSA14-44的编码区序列与人类及鼠源RAS同源基因家族核酸序列达到85%以上的同源性;RSA14-44蛋白没有典型的跨膜结构域,也没有典型的N末端信号肽;与人类RhoA蛋白序列达到了89%的同源且具有Rho家族成员的GAAX盒和p-loop结构的基序特征;RSA14-44蛋白大部分氨基酸序列与Rho家族7个已知结构域高度同源;RSA14-44基因真核表达定位于细胞质。结论：RSA14-44基因真核表达定位于CHO-K1细胞质,;编码蛋白质与Rho家族同源性高,为进一步研究其生物学功能提供参考。     

卷曲蛋白受体与肿瘤的研究进展

Wnt基因超家族编码一系列分泌型糖蛋白,由Wnt蛋白介导的Wnt信号通路是一个进化上高度保守的信号通路,在胚胎发育、细胞生长、干细胞增殖中发挥重要的作用.此外研究发现Wnt通路在癌症发生发展中也起着重要作用.Wnt信号配体-Frizzled受体,由Frizzled基因编码,属于7次跨膜蛋白受体家族,与G蛋白偶联受体的结构相似,目前研究发现,从无脊椎动物到脊椎动物至少有10个家族成员,其在心血管系统和其他器官系统中广泛表达.基于这些研究基础,本文将重点阐述Frizzled受体蛋白与疾病的可能或潜在的关系.由于Frizzled受体蛋白在疾病当中发挥了重要作用,并且在癌症中起着分子靶点的作用,我们有理由相信,Frizzled受体将成为一个有效的肿瘤治疗的分子靶点.     

The LRIG gene family has three vertebrate paralogs widely expressed in human and mouse tissues and a homolog in Ascidiacea

Human LRIG1 (formerly LIG1), human LRIG2, and mouse Lrig1 (also known as Lig-1) encode integral membrane proteins. The human genes are located at chromosomes 3p14 and 1p13, which are regions frequently deleted in human cancers. We have searched for additional members of the LRIG family and by molecular cloning identified human LRIG3 and its mouse ortholog Lrig3. Human LRIG3 is located at chromosome 12q13. In silico analysis of public databases revealed a mouse Lrig2 mRNA, three LRIG homologs in the puffer fish Fugu rubripes, and one LRIG homolog in the ascidian tunicate Ciona intestinalis. The human and mouse LRIG polypeptides have the same predicted domain organization: a signal peptide, 15 tandem leucine-rich repeats with cysteine-rich N- and C-flanking domains, three immunoglobulin-like domains, a transmembrane domain, and a cytoplasmic tail. The extracellular part--especially the IgC2.2 domain, the transmembrane domain, and the membrane-proximal part of the cytoplasmic tail--are the most conserved regions. Northern blot analysis and real-time RT-PCR revealed that the three LRIG paralogs are widely expressed in human and mouse tissues. In conclusion, the LRIG gene family was found to have three widely expressed mammalian paralogs, corresponding orthologs in fish, and a homolog in Ascidiacea.     

Leucine-rich Repeat and Immunoglobulin Domain-containing Protein-1 (Lrig1) Negative Regulatory Action toward ErbB Receptor Tyrosine Kinases Is Opposed by Leucine-rich Repeat and Immunoglobulin Domain-containing Protein 3 (Lrig3)

Lrig1 is the founding member of the Lrig family of transmembrane leucine-rich repeat proteins, which also includes Lrig2 and Lrig3. Lrig1 is a negative regulator of oncogenic receptor tyrosine kinases, including ErbB and Met receptors, and promotes receptor degradation. Lrig1 has recently emerged as both a tumor suppressor and a key regulator of epidermal and epithelial stem cell quiescence. Despite this, little is known of the mechanisms by which Lrig1 is regulated. Lrig3 was recently reported to increase ErbB receptor expression suggesting that it may function in a manner opposite to Lrig1. In this study, we explore the interaction between Lrig1 and Lrig3 and demonstrate that Lrig1 and Lrig3 functionally oppose one another. Lrig3 opposes Lrig1 negative regulatory activity and stabilizes ErbB receptors. Conversely, Lrig1 destabilizes Lrig3, limiting Lrig3''s positive effects on receptors and identifying Lrig3 as a new target of Lrig1. These studies provide new insight into the regulation of Lrig1 and uncover a complex cross-talk between Lrig1 and Lrig3.     

In Vivo Analysis of Lrig Genes Reveals Redundant and Independent Functions in the Inner Ear

Lrig proteins are conserved transmembrane proteins that modulate a variety of signaling pathways from worm to humans. In mammals, there are three family members – Lrig1, Lrig2, and Lrig3 – that are defined by closely related extracellular domains with a similar arrangement of leucine rich repeats and immunoglobulin domains. However, the intracellular domains show little homology. Lrig1 inhibits EGF signaling through internalization and degradation of ErbB receptors. Although Lrig3 can also bind ErbB receptors in vitro, it is unclear whether Lrig2 and Lrig3 exhibit similar functions to Lrig1. To gain insights into Lrig gene functions in vivo, we compared the expression and function of the Lrigs in the inner ear, which offers a sensitive system for detecting effects on morphogenesis and function. We find that all three family members are expressed in the inner ear throughout development, with Lrig1 and Lrig3 restricted to subsets of cells and Lrig2 expressed more broadly. Lrig1 and Lrig3 overlap prominently in the developing vestibular apparatus and simultaneous removal of both genes disrupts inner ear morphogenesis. This suggests that these two family members act redundantly in the otic epithelium. In contrast, although Lrig1 and Lrig2 are frequently co-expressed, Lrig1^−/−;Lrig2^−/− double mutant ears show no enhanced structural abnormalities. At later stages, Lrig1 expression is sustained in non-sensory tissues, whereas Lrig2 levels are enhanced in neurons and sensory epithelia. Consistent with these distinct expression patterns, Lrig1 and Lrig2 mutant mice exhibit different forms of impaired auditory responsiveness. Notably, Lrig1^−/−;Lrig2^−/− double mutant mice display vestibular deficits and suffer from a more severe auditory defect that is accompanied by a cochlear innervation phenotype not present in single mutants. Thus, Lrig genes appear to act both redundantly and independently, with Lrig2 emerging as the most functionally distinct family member.     

Identification of a member of mouse semaphorin family

Grasshopper semaphorin I (Sema I) and its related proteins, chick collapsin and mouse Sema III contribute to the axon guidance by their repellent actions [5,9,12]. We have identified a member of semaphorin gene family from the mouse brain and named it M-Sema F. The N-terminal encodes a semaphorin domain that is similar between Sema I–III [6] followed by a single putative immunoglobulin-like domain, a transmembrane domain, and a proline-rich intracellular domain. M-Sema F mRNA is expressed widely in the nervous tissues during development. These suggest that M-Sema F is a transmembrane member of the semaphorin family of the vertebrate which may function in the developing neuronal network.     

拟南芥和水稻SET结构域基因家族全基因组鉴定、分类和表达

ET(Su(var), Enhancer of zeste (E(z)), and Trithorax)结构域基因家族是一组含有保守SET结构域的蛋白的统称, 它们参与蛋白甲基化, 影响染色体结构, 并且调控基因表达, 在植物发育中起着重要的作用。分析拟南芥和水稻中SET结构域基因家族进化关系, 对研究这一基因家族中各成员的功能有着重要的意义。我们系统地鉴定了47个拟南芥(Arabidopsis thaliana)和43个水稻(Orysa sativa japonica cultivar Nipponbare)的SET结构域基因, 染色体定位和基因复制分析表明SET结构域基因扩增是由片段复制和反转录引起的, 根据这些结构域差异和系统发育分析把拟南芥和水稻的SET结构域基因划分成5个亚家族。通过分析SET结构域基因家族在拟南芥和水稻各个发育阶段的表达谱, 发现SET结构域基因绝大部分至少在一个组织中表达; 大部分在花和花粉中高表达; 一些SET结构域基因在某些组织中有特异的表达模式, 表明与组织发育有密切的关系。在拟南芥和水稻中分别找到了4个差异表达基因。拟南芥4个差异基因都在花粉管高表达, 水稻4个差异基因有3个在雄性花蕊中高表达, 另一个在幼穗中高表达。     

甘蔗细胞色素C基因的电子克隆与分析

以甘蔗类似细胞色素C的EST序列CF576943.1为探针,通过电子克隆技术获得了甘蔗细胞色素C基因（Cytochrome C,Cyt C）的一条cDNA全长序列,命名为ScCyt C.用生物信息学方法对该基因氨基酸序列与组成、亚细胞定位、跨膜区与信号肽、疏水性/亲水性、蛋白质二、三级结构以及功能等进行分析与预测.结果表明：Cyt C基因全长1 073 bp,编码112个氨基酸,该基因位于细胞质,为非分泌型蛋白,无规卷曲为主要二级结构原件,含有1个保守功能域,主要功能为翻译并且在不同植物中具有高度保守性.电子表达分析结果显示,该基因在甘蔗各个组织均有表达,其中在茎和根中的表达量比其他组织类型中表达量高,此外,该基因的表达可能受到低温的调控.为甘蔗细胞色素C基因的结构及其功能的研究奠定了一定的基础.     

甘蔗天冬氨酰半醛脱氢酶基因的电子克隆与分析

应用电子克隆技术,以水稻EF576477序列为探针,获得了甘蔗天冬氨酰半醛脱氢酶基因（aspartate．semialdehydedehy—drogenase,ASADH）的一条cDNA全长序列,命名为ScASADH。采用生物信息学方法,对该基因编码蛋白从氨基酸组成、理化性质、亚细胞定位、跨膜结构域、疏水性／亲水性、高级结构及功能域等方面进行预测和分析。结果表明：该基因全长1711bp,包含一个1128bp的开放阅读框,编码375个氨基酸,该基因编码蛋白定位于细胞核,为可溶性蛋白,存在信号肽,二级结构原件多为无规卷曲,含有多个保守功能域,主要功能为翻译。电子表达分析结果显示,该基因在甘蔗根尖、幼苗、花序、叶片和茎中组成型表达,其中在茎中的表达量比其他组织类型中表达量高。该基因的表达受葡萄糖杆菌和赤腐病菌的调控。     

Molecular cloning of MIS, a myeloid inhibitory siglec, that binds protein-tyrosine phosphatases SHP-1 and SHP-2

We describe the molecular cloning and characterization of a novel myeloid inhibitory siglec, MIS, that belongs to the family of sialic acid-binding immunoglobulin-like lectins. A full-length MIS cDNA was obtained from murine bone marrow cells. MIS is predicted to contain an extracellular region comprising three immunoglobulin-like domains (V-set amino-terminal domain followed by two C-set domains), a transmembrane domain and a cytoplasmic tail with two immunoreceptor tyrosine-based inhibitory motif (ITIM)-like sequences. The closest relative of MIS in the siglec family is human siglec 8. Extracellular regions of these two siglecs share 47% identity at the amino acid level. Southern blot analysis suggests the presence of one MIS gene. MIS is expressed in the spleen, liver, heart, kidney, lung and testis tissues. Several isoforms of MIS protein exist due to the alternative splicing. In a human promonocyte cell line, MIS was able to bind Src homology 2-containing protein-tyrosine phosphatases, SHP-1 and SHP-2. This binding was mediated by the membrane-proximal ITIM of MIS. Moreover, MIS exerted an inhibitory effect on FcgammaRI receptor-induced calcium mobilization. These data suggest that MIS can play an inhibitory role through its ITIM sequences.