文昌鱼分类学研究及展望

文昌鱼是最接近脊椎动物直接祖先的现生动物,在脊椎动物起源与演化研究中占有极其重要的位置。近年来,对文昌鱼的研究已引起越来越多的科学家的兴趣,然而作为生命科学研究的重要基础,这类动物的分类学研究相对滞后。依据已有的中国文昌鱼资源调查资料,中国沿海文昌鱼的分布应当十分广泛,即只要有适合文昌鱼栖息的沙滩,均有文昌鱼分布的可能。根据目前的分类学研究成果和动物命名法中的优先权原则,建议将产于青岛等地的文昌鱼种名Brnachiostoma belcheri tsingtauense订正为B.japonicus,南方的文昌鱼保留其原种名B.belcheri。由此,目前分布在中国沿海的鳃口文昌鱼属(Branchiostoma)至少有2种,侧殖文昌鱼属(Epigonichthys)有1～3种,漂浮文昌鱼(Amphioxides pelagicus)1种。DNA分子标记技术在文昌鱼分类学研究中将会发挥更大的作用。     

文昌鱼—研究脊柱动物起源和进化的模式动物

长久以来,文昌鱼一直被认为和生活在约5亿年前的脊椎动物的直接祖先相似。由于文昌鱼在进化上的重要性,它在动物学研究史上发挥着关键作用,近100多年来,文昌鱼作为研究对象曾数次受到动物学界青睐或冷落,大约10年前,随着分子生物学技术应用于文昌鱼研究,又激发了动物学家对文昌鱼的研究兴趣,又一次出现在文昌鱼研究的高潮,并且一直持续至今,分子生物学研究结果表明,文昌鱼样生物可能是环节动物样动物和最早的脊椎动物之间的进化中间体,因此,文昌鱼在动物学研究史上好像绕了个大圈又回到了原处,在被忽视一段时间之后,又重新占据脊椎动物起源和进化研究中心舞台的位置,成为研究脊椎动物起源和进化的模式动物。     

文昌鱼抗金葡菌感染差减文库的构建及免疫相关基因分析

适应性免疫的起源一直是免疫学研究的关键问题.文昌鱼被认为是最接近于脊椎动物的祖先 自从被发现以来一直是研究脊椎动物起源与进化机制的经典模式动物.为了在文昌鱼中寻找适应性免疫系统的分子证据,采用金黄色葡萄球菌感染文昌鱼以调查免疫的起源.应用抑制性差减杂交(SSH)技术,通过对差减文库克隆序列的测定,共获得588个表达序列标签(EST).对这些EST进行生物信息学分析和进一步功能分类,发现了一些免疫上调基因,如免疫调控基因、凋亡相关基因、细胞黏附相关基因、转录相关基因、信号传导相关基因等,以及一些非免疫相关基因;这些基因在文昌鱼中绝大多数为首次报道.金黄色葡萄球菌差减文库的成功构建,为调查文昌鱼抗细菌感染的分子事件提供了重要线索,对于这些新发现基因的进一步研究将有助于深入了解免疫系统起源与进化的机制.     

我国文昌鱼研究50年

文昌鱼是研究脊椎动物起源与进化十分珍贵的模式动物。近50年来,我国在文昌鱼生物学研究方面取得很大进展。本文系统介绍了我国学者在文昌鱼发育、免疫和进化方面所取得的研究成果。     

文昌鱼转铁蛋白及转铁蛋白的分子进化

从厦门文昌鱼分离纯化了文昌鱼转铁蛋白,物化性质与青岛文昌鱼转铁蛋白相同,其单体和二聚体的分子量分别为２６ｋＤ和５２ｋＤ,是一分子量约为脊椎动物转铁蛋白１／４的糖蛋白,测定了文昌鱼转铁蛋白完整分子和其Ｃ端分子片段的部分氨基酸序列并分析 一人务清失蛋白氨基酸序列的同源性。发现用文昌鱼转铁蛋白序列可将人血清转铁蛋白序列划分成粗略相等的４个片段,文是鱼转铁蛋白与每一片段及人血清转铁蛋白的４个片段之间存在明     

文昌鱼AmphiRab23b基因的克隆、进化分析及表达模式

Hedgehog信号通路在胚胎发育过程中发挥着重要作用, 同时与多种肿瘤的发生密切相关。Rab23蛋白在Hedgehog信号通路中扮演着十分重要的角色。目前关于文昌鱼Rab23同源基因的研究仅局限于佛罗里达文昌鱼(Branchiostoma floridae)基因组中的注释。文章首次克隆了中国文昌鱼(Branchiostoma belcheri) Rab23b基因 (AmphiRab23b)cDNA全序列 , 对其演译的蛋白序列进行了序列比对、进化树分析以及基因时空表达分析。研究结果显示, 文昌鱼AmphiRab23b基因的 cDNA总长为2 062 bp (包括UTR区), 其中开放阅读框 (Open reading frame, ORF) 714 bp, 编码237个氨基酸; 虽然在进化树中不属于脊椎动物Rab23进化支, 但是AmphiRab23具有保守的Rab23_lke结构域, 暗示该基因在进化过程中可能在功能上是保守的。时空表达的研究结果进一步显示, AmphiRab23b基因在胚胎发育中的神经板和消化道中表达, 与其脊椎动物同源基因的表达模式相似, 这说明该基因可能对文昌鱼神经系统和消化道的发育有重要作用。     

文昌鱼左右体轴建立机制的研究进展

两侧对称动物左右体轴建立机制研究是发育生物学领域重要的基础科学问题之一。文昌鱼(amphioxus)由于其特殊的进化地位以及与脊椎动物相似的胚胎发育模式和身体构筑方式,是研究动物左右体轴建立机制的理想模式物种。近年来随着文昌鱼室内全人工繁育技术、高效显微注射技术和基因敲除技术的建立,国内外学者在左右体轴建立机制研究上取得了丰硕的成果。本文从文昌鱼胚胎左右不对称发育特点出发,总结了近期文昌鱼左右体轴建立方面取得的研究进展,并提出了文昌鱼左右体轴调控网络图:纤毛运动导致Hh蛋白在文昌鱼中不对称分布(L相似文献   

文昌鱼AmphiDC-like的全长cDNA克隆及其表达模式分析

本文旨在克隆文昌鱼树突样蛋白基因AmphiDC-like,采用实时 PCR法和原位分子杂交法对其表达模式进行分析.从文昌鱼神经胚cDNA文库测序得到的ESTs中筛选得到该基因片段,通过引物步移直接测序的方法,克隆得到其cDNA全长序列,对其推测的氨基酸序列进行同源性分析发现,该基因产物具有树突样细胞蛋白共有的保守区,并与多种生物的树突状细胞蛋白具有高度同源性,其中与脊椎动物的树突样细胞蛋白同源性较高. 实时 PCR结果显示, AmphiDC-like在文昌鱼受副溶血性弧菌(Vibrio parahemolyticus,Vp)感染后6 h到48 h表达均上调,并通过原位分子杂交技术观察到,该基因在文昌鱼鳃和消化道中有表达,为深入研究其功能奠定了基础.     

文昌鱼的实验室繁育及子二代获得

文昌鱼特殊的进化地位、简单的器官系统和终生透明的躯体等特征，使其很有希望成为一个新型实验室模式动物。要实现文昌鱼的模式动物化，实验室内全人工条件下繁殖是关键的第一步。为此，我们于2003年9月和2004年4月两次采集产于厦门海域的2种文昌鱼，开展实验室内养殖研究。经过3年多的持续实验室养殖，继2005年夏季于实验室内繁殖出子一代文昌鱼后，又在2006年夏季成功获得了这两种文昌鱼的子二代，初步实现文昌鱼在实验室内的全人工养殖。对子代文昌鱼养殖的初步观察发现，不同水温对生长发育速度有一定影响，提示有可能通过水温控制实现文昌鱼一年多次产卵的目的。目前这两种文昌鱼子二代幼体已完成变态，进入亚成体生长发育阶段，其体长分别已达14.6 mm(日本文昌鱼Branchiostoma japonicum)和6.5 mm(白氏文昌鱼B. belcheri)。     

文昌鱼的实验室繁育及子二代获得

文昌鱼特殊的进化地位、简单的器官系统和终生透明的躯体等特征,使其很有希望成为一个新型实验室模式动物。要实现文昌鱼的模式动物化,实验室内全人工条件下繁殖是关键的第一步。为此,我们于2003年9月和2004年4月两次采集产于厦门海域的2种文昌鱼,开展实验室内养殖研究。经过3年多的持续实验室养殖,继2005年夏季于实验室内繁殖出子一代文昌鱼后,又在2006年夏季成功获得了这两种文昌鱼的子二代,初步实现文昌鱼在实验室内的全人工养殖。对子代文昌鱼养殖的初步观察发现,不同水温对生长发育速度有一定影响,提示有可能通过水温控制实现文昌鱼一年多次产卵的目的。目前这两种文昌鱼子二代幼体已完成变态,进入亚成体生长发育阶段,其体长分别已达14.6mm(日本文昌鱼Branchiostomajaponicum)和6.5mm(白氏文昌鱼B.belcheri)。     

Cis-regulation and conserved non-coding elements in amphioxus

The origin of the vertebrates was a major event in the evolution of morphological diversity and the genetic mechanisms responsible for this diversity, once purely theoretical, can now be approached experimentally in the genome era. With a prototypical chordate genome, vertebrate-like development and simple morphology, amphioxus provides the appropriate model for investigating the origin of the vertebrates. Comparative genomics is revealing that both conservation and divergence of genes and cis-regulatory elements involved in developmental regulatory networks are required to shape different animal body plans. This article reviews the cis-regulatory studies performed in amphioxus, the discovery of conserved non-coding elements (CNEs) across the metazoans and the examination of amphioxus CNEs. Emerging ideas on the evolution of CNEs after large-scale genome duplication events and the state of cephalochordate genomics are also discussed.     

The basal chordate amphioxus as a simple model for elucidating developmental mechanisms in vertebrates

This review examines the basal chordate, amphioxus, as a simple model for providing insights into the development and evolution of the vertebrates, with which it shares many features, including a pharynx perforated with gill slits, a dorsal nerve cord, segmented muscles, and a notochord. Conversely, amphioxus is simpler than vertebrates in lacking neural crest and paired cephalic sensory organs. Amphioxus embryos are less derived than those of vertebrates, because it lacks large quantities of yolk and/or extra-embryonic tissues. Embryogenesis involves only a simple folding of tissue layers. In addition, the amphioxus genome lacks the large-scale gene duplications of vertebrates. However, in spite of the comparative simplicity of amphioxus, its developmental mechanisms are proving to be highly conserved with those of vertebrates. Thus, studies of amphioxus development can shed light on similar, but more complex, development of vertebrates. Such studies are especially interesting for their insights into the genetic basis of craniofacial birth defects in humans.     

Amphioxus: a peaceful anchovy fillet to illuminate Chordate Evolution (II)

The genome of the amphioxus is on the horizon. With Linda Holland and Jeremy Gibson-Brown at the forefront, with all the amphioxus community behind, and with the Joint Genome Institute, the amphioxus genome will see the light this year, 2006. Hope that it will reflect the "prototypical" preduplicative genome of vertebrates. It may answer definitively what the human genome did not: Are we vertebrates octaploid? Will it shed light on the novelties that helped non-chordates to be chordates? And more, will amphioxus, with a simpler genome, be developed to a senior "experimental model system", allowing the testing of molecular functions in a non-duplicated genome background and allowing genetic modification to "recapitulate" evolution? Thanks to an outstanding collaboration between labs, the laboratory culture of amphioxus is underway after years of hard work in the field. 2007 looks promising for amphioxus research.     

Free amino acids in the nervous system of the amphioxus Branchiostoma lanceolatum. A comparative study

The cephalochordate amphioxus is the closest invertebrate relative to vertebrates. In this study, using HPLC technique, free L-amino acids (L-AAs) and D-aspartic acid (D-Asp) have been detected in the nervous system of the amphioxus Branchiostoma lanceolatum. Among other amino acids glutamate, aspartate, glycine, alanine and serine are the amino acids found at the greatest concentrations. As it occurs in the nervous system of other animal phyla, glutamate (L-Glu) and aspartate (L-Asp) are present at very high concentrations in the amphioxus nervous system compared to other amino acids, whereas the concentration of taurine and gamma-aminobutyric acid (GABA) is very low. Interestingly, as it is the case in vertebrates, D-aspartic acid is present as an endogenous compound in amphioxus nervous tissues. The physiological function of excitatory amino acids, and D-aspartate in particular, are discussed in terms of evolution of the nervous system under an Evo-fun (Evolution of function) perspective.     

Consecutive Spawnings of Chinese Amphioxus,Branchiostoma belcheri,in Captivity

Cephalochordate amphioxus is a promising model animal for studying the evolutionary and developmental mechanisms of vertebrates because its unique phylogenetic position, simple body plan and sequenced genome. However, one major drawback for using amphioxus as a model organism is the restricted supply of living embryos since they are available only during spawning season that varies from a couple of days to several months according to species. Therefore we are aiming to develop methods for obtaining viable amphioxus embryos in non-spawning season. In the current study, we found that Branchiostoma belcheri could develop their gonads and spawn consecutively in the laboratory when cultured in a low density at a high temperature (25–28°C) supplied with sufficient food and proper cleanness. Among the approximate 150 observed animals, which spawned spontaneously between November and December 2011, 10% have spawned twice, 10% three times, and 80% four times, through April 2012. The quality and quantity of the gametes reproduced in the consecutive spawning have no obvious difference with those spawned once naturally. Spawning intervals varied dramatically both among different animals (from 1 to 5 months) and between intervals of a single individual (from 27 to 74 days for one animal). In summary, we developed a method with which, for the first time, consecutive spawnings of amphioxus in captivity can be achieved. This has practical implications for the cultivation of other amphioxus species, and eventually will greatly promote the utilization of amphioxus as a model system.     

Identification and characterisation of five novel miniature inverted-repeat transposable elements (MITEs) in amphioxus (Branchiostoma floridae)

As the sister group to vertebrates, amphioxus is consistently used as a model of genome evolution for understanding the invertebrate/vertebrate transition. The amphioxus genome has not undergone massive duplications like those in the vertebrates or disruptive rearrangements like in the genome of Ciona, a urochordate, making it an ideal evolutionary model. Transposable elements have been linked to many genomic evolutionary changes including increased genome size, modified gene expression, massive gene rearrangements, and possibly intron evolution. Despite their importance in genome evolution, few previous examples of transposable elements have been identified in amphioxus. We report five novel Miniature Inverted-repeat Transposable Elements (MITEs) identified by an analysis of amphioxus DNA sequence, which we have named LanceleTn-1, LanceleTn-2, LanceleTn-3a, LanceleTn-3b and LanceleTn-4. Several of the LanceleTn elements were identified in the amphioxus ParaHox cluster, and we suggest these have had important implications for the evolution of this highly conserved gene cluster. The estimated high copy numbers of these elements implies that MITEs are probably the most abundant type of mobile element in amphioxus, and are thus likely to have been of fundamental importance in shaping the evolution of the amphioxus genome.     

Genomics reveal ancient forms of stanniocalcin in amphioxus and tunicate

Stanniocalcin (STC) is present throughout vertebrates, including humans, but a structure for STC has not been identified in animals that evolved before bony fish. The origin of this pleiotropic hormone known to regulate calcium is not clear. In the present study, we have cloned three stanniocalcins from two invertebrates, the tunicate Ciona intestinalis and the amphioxus Branchiostoma floridae. Both species are protochordates with the tunicates as the closest living relatives to vertebrates. Amphioxus are basal to both tunicates and vertebrates. The genes and predicted proteins of tunicate and amphioxus share several key structural features found in all previously described homologs. Both the invertebrate and vertebrate genes have four conserved exons. The predicted length of the single pro-STC in Ciona is 237 amino acids and the two pro-hormones in amphioxus are 207 and 210 residues, which is shorter than human pro-STCs at 247 and 302 residues due to expansion of the C-terminal region in vertebrate forms. The conserved pattern of 10 cysteines in all chordate STCs is crucial for identification as amphioxus and tunicate amino acids are only 14-23% identical with human STC1 and STC2. The 11th cysteine, which is the cysteine shown to form a homodimer in vertebrates, is present only in amphioxus STCa, but not in amphioxus STCb or tunicate STC, suggesting the latter two are monomers. The expression of stanniocalcin in Ciona is widespread as shown by RT-PCR and by quantitative PCR. The latter method shows that the highest amount of STC mRNA is in the heart with lower amounts in the neural complex, branchial basket, and endostyle. A widespread distribution is present also in mammals and fish for both STC1 and STC2. Stanniocalcin is a presumptive regulator of calcium in both Ciona and amphioxus, although the structure of a STC receptor remains to be identified in any organism. Our data suggest that amphioxus STCa is most similar to the common ancestor of vertebrate STCs because it has an 11th cysteine necessary for dimerization, an N-glycosylation motif, although not the canonical one in vertebrate STCs, and similar gene organization. Tunicate and amphioxus STCs are more similar in structure to vertebrate STC1 than to vertebrate STC2. The unique features of STC2, including 14 instead of 11 cysteines and a cluster of histidines in the C-terminal region, appear to be found exclusively in vertebrates.