Sox1基因在爪蟾早期发育中的时空表达图式

克隆了非洲爪蟾的Sox1基因并研究了它在非洲爪蟾早期发育过程中的时空表达图式,比较了Sox1—3基因在发育的脑和眼中的表达图式。序列比对分析显示Sox1—3蛋白在其HMG框结构域具有高度的保守性。通过RT-PCR方法分析了Sox1基因在爪蟾早期不同发育时段的表达情况,结果显示Sox1基因从未受精卵到尾芽期均有表达,但表达强度有所差异。原位杂交结果显示,在早期卵裂阶段和囊胚期,Sox1基因主要在动物极表达;从神经板期开始,Sox1基因主要在中枢神经系统和眼原基中表达。在蝌蚪期,Sox1与Sox2、Sox3在脑部和眼睛的表达区域有所不同。对于爪蟾Sox1基因时空表达图式的研究将有助于阐明SoxB1基因家族在脊椎动物神经系统发生过程中的作用。     

东北虎中五个Sox基因的保守区序列分析(英文)

在哺乳动物中,Y染色体决定着雄性性别,这是由于在其短臂上存在一个编码睾丸决定因子 (TDF) 的基因。1990年,人们克隆了睾丸决定因子基因并命名为SRY。序列分析表明SRY基因中存在一个保守区,与染色体高迁移率组 (HMG) 蛋白质上的DNA结合结构域具有一定的相似性。基于HMG基序的保守性人们发现了一个新的基因家族Sox基因家族。凡是在HMG区域与SRY基因具有50%以上相似性的基因被称为Sox基因。Sox基因在早期胚胎发育过程中参与多种发育途径,具有重要的作用。参与诸如性别决定、骨组织的发育、血细胞生成过程、神经系统的发育、晶状体的发育等多种早期胚胎发育过程。 虎(Panthera tigris)作为世界上最濒危的兽类之一,东北虎(Panthera tigris altaica Temminck)是其中的一个亚种,被列为一类珍稀动物。本文对其发育基因家族—SOX基因进行了研究。 利用肌肉组织为材料制备基因组DNA, 应用特异于HMG-box区域的兼并引物, 扩增了东北虎的SOX基因。在扩增产物中发现一条220bp的扩增带。经过克隆与序列测定和同源性检索,发现5个基因片段(Fig.1&2)。其与人SRY基因的相似性分别为75%、56%、51%、67% 和48%;与小鼠Sry基因的相似性分别为73%、54%、57%、66% 和 48% (Table 1)。因此这5个DNA片段为东北虎的5个Sox基因片     

转录因子Sox2的研究进展

转录因子Sox2是sox基因家族的一个成员,由于它在早期胚胎发生、神经分化和晶状体发育等多种重要的发育事件中都起着关键的作用,从而引起了越来越广泛的关注。本文就小鼠sox2基因的研究进展作一综述。     

小鼠内耳发育核心转录因子的保守性及调控网络的比较分析

在脊椎动物内耳发育中, Six1、Six4、Pax2、Pax8、Foxi1、Dlx5、Gbx2、Irx2/3、Msx1等基因作为核心调控基因参与听基板的诱导过程。文章通过生物信息学方法, 对小鼠内耳发育的核心转录因子进行保守性分析并研究其相互调控关系, 得到小鼠内耳发育过程中核心转录因子的基因调控网络。与文献中已知的小鼠内耳发育基因调控关系相比, Pax2、Pax8、Foxi1、Dlx5基因在内耳发育中仍然起主要调控者的角色, Six1则处于被多个转录因子调节的地位, Gbx2、Irx2/3、Msx1在调控网络中也起到重要作用。对出现的差异进行了合理的分析, 同时结合构建的调控网络预测了可能存在的Msx1对Six1、Gbx2的调控作用。序列预测结果也发现了一些新的调控关系, 所涉及的转录因子包括Sox5、Lhx2、Rax、Otx1、Otx2、Pitx1、Pitx2、Nkx2-5、Irx4、Irx6、Dlx2、Hmx1/2/3、Pou4f3、Pax4、Tlx2。文章为深入了解内耳发育调控机制提供了基础信息。     

Sox基因家族功能的研究进展

Sox（Sry-related high mobility group box）基因是由众多具有HMG-box保守基序构成的超基因家族,是与位于雄性动物Y染色体上的Sry基因同源的家族基因,在很多动物中都有表达。由于其编码的蛋白质具有结合DNA的能力,因而认为Sox基因家族是一类重要的转录调控因子。在个体发育过程中,Sox基因广泛参与了动物的早期胚胎发育、性别决定和分化、神经系统发育、软骨及多种组织器官的形成,具有重要的生物学功能。主要对Sox家族基因的功能及其研究进展进行简要的综述。     

马氏珠母贝Sox11基因的克隆及时序表达模式分析

为探究Sox(SRY-related HMG-box genes)基因家族在马氏珠母贝个体发育及性别分化中的作用, 研究首先利用兼并引物从马氏珠母贝基因组中克隆到一个HMG框(high mobility group box), 利用RACE-PCR技术从SMART cDNA文库中克隆到一个Sox基因的cDNA全长, 通过Clustal X和MEGA 4软件对该序列进行比对分析并构建系统进化树; 通过荧光定量PCR技术对该基因在不同组织及发育不同时期性腺中的表达情况进行分析。结果显示, 马氏珠母贝该Sox基因的cDNA全长为1579 bp, 其中开放阅读框(ORF)为1008 bp, 编码336个氨基酸, 5'非编码区为126 bp, 3'非编码区为445 bp。同源性分析表明, 马氏珠母贝Sox基因与太平洋牡蛎Sox11基因的同源性(Identity)最高, 为80%, 故命名为pmSox11; 系统进化树分析也显示pmSox11与太平洋牡蛎Sox11基因的亲缘关系最近。荧光定量PCR分析组织表达特异性显示, pmSox11在马氏珠母贝神经节分布较多的组织如外套膜、鳃、足、消化盲囊等大量表达, 在神经节相对较少的闭壳肌和卵巢中表达量较少; 时序表达图谱显示, pmSox11在3月龄幼贝性腺和1年龄发育早期精巢中表达量最高, 在2年龄成熟精巢和2年龄性转换性腺中表达量降低, 而在2年龄卵巢中表达量最低。研究表明, pmSox11基因可能在马氏珠母贝早期神经系统发育和性别发育的调控方面起到重要作用。     

中国大鲵Sox2基因全长cDNA序列的克隆及组织表达分析

Sox2是Sox转录因子超家族B1亚族成员之一,对胚胎发育和神经细胞的维持有重要作用。本研究以中国大鲵(Andrias davidianus)性腺组织为材料,通过RT-PCR和RACE方法成功克隆到中国大鲵Sox2基因全长2 233 bp c DNA。系统进化树分析表明,中国大鲵与红腹蝾螈、火焰蝾螈及六角恐龙聚成一支,这与其在两栖类中分化较早相一致。实时荧光定量PCR检测Sox2基因在中国大鲵组织表达结果显示,其在性腺中的表达量最高,肾脏中次之,在肠道中几乎不表达,表明Sox2基因可能在大鲵性腺发育和生长发育过程中发挥了重要作用。这为今后进一步研究Sox2基因在大鲵性别分化中的功能提供了参考。     

Sox9基因的结构、功能及进化

Sox9基因是继SRY基因之后人们发现的第一个具有内含子的Sox家族成员。在Genebank中可查的Sox9基因序列共671个,广泛分布于哺乳动物和鸟类直至水母等低等动物。近年来的研究发现,Sox9基因的功能呈现多样性,该基因在发育过程中参与性别的决定、软骨的形成以及神经系统和胰腺的形成,另外Sox9基因在肿瘤发生中也有一定的作用。本文从Sox9基因的结构、功能以及进化等方面进行了综述,为进一步研究Sox9基因提供参考资料。     

翘嘴鲌Sox9基因的克隆及CpG岛甲基化与基因表达的关系

为了揭示翘嘴鲌(Culter alburnus)性别决定与分化的作用机制, 进而更好地发展性别控制育种技术, 研究重点分析了Sox9基因在翘嘴鲌性腺分化过程中的作用。通过RT-PCR和RACE方法获得了翘嘴鲌2个旁系同源基因Sox9a和Sox9b的cDNA序列: Sox9a全长1642 bp, 编码458个氨基酸; Sox9b全长1673 bp, 编码456个氨基酸。序列分析表明两者相似度达到73.95%, 编码HMG盒区域极其保守。蛋白质次级结构预测显示Sox9a和Sox9b除了保守的HMG盒结构域外, 还存在2个核定位信号; 两者的三维结构都存在多个螺旋结构。系统进化树分析发现翘嘴鲌Sox9a与罗非鱼关系最近, 但Sox9b形成单独的一支。利用实时荧光定量PCR技术分析了翘嘴鲌Sox9a和Sox9b基因在各成体组织中的表达水平, 结果显示Sox9a在脑和精巢中表达量最高, 其次是肌肉、鳍条、眼睛和卵巢, 在肾脏、脾脏、肝脏中相对较低; Sox9b只在脑、鳍条、眼睛和精巢中检测到一定水平的表达。通过重亚硫酸氢盐DNA测序方法分析了翘嘴鲌性腺组织Sox9a启动子CpG岛甲基化修饰模式, 结果显示在精巢中CG位点几乎不发生甲基化, 然而卵巢中的甲基化程度非常高。这些结果表明启动子CpG甲基化可以调控Sox9a的性别异形表达, 表观遗传修饰在翘嘴鲌性腺发育过程中可能具有重要的生物学功能。     

Dual role for Sox2 in specification of sensory competence and regulation of Atoh1 function

The formation of inner ear sensory epithelia is believed to occur in two steps, initial specification of sensory competent (prosensory) regions followed by determination of specific cell‐types, such as hair cells (HCs) and supporting cells. However, studies in which the HC determination factor Atoh1 was ectopically expressed in nonprosensory regions indicated that expression of Atoh1 alone is sufficient to induce HC formation suggesting that prosensory formation may not be a prerequisite for HC development. To test this hypothesis, interactions between Sox2 and Atoh1, which are required for prosensory and HC formation respectively, were examined. Forced expression of Atoh1 in nonprosensory cells resulted in transient expression of Sox2 prior to HC formation, suggesting that expression of Sox2 is required for formation of ectopic HCs. Moreover, Atoh1 overexpression failed to induce HC formation in Sox2 mutants, confirming that Sox2 is required for prosensory competence. To determine whether expression of Sox2 alone is sufficient to induce prosensory identity, Sox2 was transiently activated in a manner that mimicked endogenous expression. Following transient Sox2 activation, nonprosensory cells developed as HCs, a result that was never observed in response to persistent expression of Sox2. These results, suggest a dual role for Sox2 in inner ear formation. Initially, Sox2 is required to specify prosensory competence, but subsequent down‐regulation of Sox2 must occur to allow Atoh1 expression, most likely through a direct interaction with the Atoh1 promoter. These results implicate Sox2‐mediated changes in prosensory cells as an essential step in their ability to develop as HCs. © 2016 Wiley Periodicals, Inc. Develop Neurobiol 77: 3–13, 2017     

Sox2 is required for maintenance and regeneration, but not initial development, of hair cells in the zebrafish inner ear

Sox2 has been variously implicated in maintenance of pluripotent stem cells or, alternatively, early stages of cell differentiation, depending on context. In the developing inner ear, Sox2 initially marks all cells in the nascent sensory epithelium and, in mouse, is required for sensory epithelium formation. Sox2 is eventually downregulated in hair cells but is maintained in support cells, the functional significance of which is unknown. Here we describe regulation and function of sox2 in the zebrafish inner ear. Expression of sox2 begins after the onset of sensory epithelium development and is regulated by Atoh1a/b, Fgf and Notch. Knockdown of sox2 does not prevent hair cell production, but the rate of accumulation is reduced due to sporadic death of differentiated hair cells. We next tested the capacity for hair cell regeneration following laser ablation of mature brn3c:gfp-labeled hair cells. In control embryos, regeneration of lost hair cells begins by 12 h post-ablation and involves transdifferentiation of support cells rather than asymmetric cell division. In contrast, regeneration does not occur in sox2-depleted embryos. These data show that zebrafish sox2 is required for hair cell survival, as well as for transdifferentiation of support cells into hair cells during regeneration.     

Notch signalling is needed to maintain, but not to initiate, the formation of prosensory patches in the chick inner ear

Notch signalling is well-known to mediate lateral inhibition in inner ear sensory patches, so as to generate a balanced mixture of sensory hair cells and supporting cells. Recently, however, we have found that ectopic Notch activity at an early stage can induce the formation of ectopic sensory patches. This suggests that Notch activity may have two different functions in normal ear development, acting first to promote the formation of the prosensory patches, and then later to regulate hair-cell production within the patches. The Notch ligand Serrate1 (Jag1 in mouse and humans) is expressed in the patches from an early stage and may provide Notch activation during the prosensory phase. Here, we test whether Notch signalling is actually required for prosensory patch development. When we block Notch activation in the chick embryo using the gamma-secretase inhibitor DAPT, we see a complete loss of prosensory epithelial cells in the anterior otocyst, where they are diverted into a neuroblast fate via failure of Delta1-dependent lateral inhibition. The cells of the posterior prosensory patch remain epithelial, but expression of Sox2 and Bmp4 is drastically reduced. Expression of Serrate1 here is initially almost normal, but subsequently regresses. The patches of sensory hair cells that eventually develop are few and small. We suggest that, in normal development, factors other than Notch activity initiate Serrate1 expression. Serrate1, by activating Notch, then drives the expression of Sox2 and Bmp4, as well as expression of the Serrate1 gene itself. The positive feedback maintains Notch activation and thereby preserves and perhaps extends the prosensory state, leading eventually to the development of normal sensory patches.     

The role of Paraxial Protocadherin in Xenopus otic placode development

Vertebrate inner ear develops from its rudiment, otic placode, which later forms otic vesicle and gives rise to tissues comprising the entire inner ear. Although several signaling molecules have been identified as candidates responsible for inner ear specification and patterning, many details remain elusive. Here, we report that Paraxial Protocadherin (PAPC) is required for otic vesicle formation in Xenopus embryos. PAPC is expressed strictly in presumptive otic placode and later in otic vesicle during inner ear morphogenesis. Knockdown of PAPC by dominant-negative PAPC results in the failure of otic vesicle formation and the loss of early inner ear markers Sox9 and Tbx2, suggesting the requirement of PAPC in the early stage of otic vesicle development. However, PAPC alone is not sufficient to induce otic placode formation.     

Jagged 1 regulates the restriction of Sox2 expression in the developing chicken inner ear: a mechanism for sensory organ specification

Hair cells of the inner ear sensory organs originate from progenitor cells located at specific domains of the otic vesicle: the prosensory patches. Notch signalling is necessary for sensory development and loss of function of the Notch ligand jagged 1 (Jag1, also known as serrate 1) results in impaired sensory organs. However, the underlying mechanism of Notch function is unknown. Our results show that in the chicken otic vesicle, the Sox2 expression domain initially contains the nascent patches of Jag1 expression but, later on, Sox2 is only maintained in the Jag1-positive domains. Ectopic human JAG1 (hJag1) is able to induce Sox2 expression and enlarged sensory organs. The competence to respond to hJag1, however, is confined to the regions that expressed Sox2 early in development, suggesting that hJag1 maintains Sox2 expression rather than inducing it de novo. The effect is non-cell-autonomous and requires Notch signalling. hJag1 activates Notch, induces Hes/Hey genes and endogenous Jag1 in a non-cell-autonomous manner, which is consistent with lateral induction. The effects of hJag1 are mimicked by Jag2 but not by Dl1. Sox2 is sufficient to activate the Atoh1 enhancer and to ectopically induce sensory cell fate outside neurosensory-competent domains. We suggest that the prosensory function of Jag1 resides in its ability to generate discrete domains of Notch activity that maintain Sox2 expression within restricted areas of an extended neurosensory-competent domain. This provides a mechanism to couple patterning and cell fate specification during the development of sensory organs.     

Sox2 and Fgf interact with Atoh1 to promote sensory competence throughout the zebrafish inner ear

Atoh1 is required for differentiation of sensory hair cells in the vertebrate inner ear. Moreover, misexpression of Atoh1 is sufficient to establish ectopic sensory epithelia, making Atoh1 a good candidate for gene therapy to restore hearing. However, competence to form sensory epithelia appears to be limited to discrete regions of the inner ear. To better understand the developmental factors influencing sensory-competence, we examined the effects of misexpressing atoh1a in zebrafish embryos under various developmental conditions. Activation of a heat shock-inducible transgene, hs:atoh1a, resulted in ectopic expression of early markers of sensory development within 2 h, and mature hair cells marked by brn3c:GFP began to accumulate 9 h after heat shock. The ability of atoh1a to induce ectopic sensory epithelia was maximal when activated during placodal or early otic vesicle stages but declined rapidly thereafter. At no stage was atoh1a sufficient to induce sensory development in dorsal or lateral non-sensory regions of the otic vesicle. However, co-misexpression of atoh1a with fgf3, fgf8 or sox2, genes normally acting in the same gene network with atoh1a, stimulated sensory development in all regions of the otic vesicle. Thus, expression of fgf3, fgf8 or sox2 strongly enhances competence to respond to Atoh1.