卡哈尔体——功能复杂的核内细胞器

卡哈尔体（Cajalbody,CB）的发现可以溯源至100多年前,而直到近10年的研究取得突破性进展,才逐渐揭示了其复杂的分子组成及生理功能。作为当前备受关注的核内小体之一,CB是细胞核质中处于动态变化的细胞器,广泛存在于高等真核生物的细胞核内,在电镜下表现为球形小体样结构,其中p80Coilin蛋白是CB最为明确的特异性标志物。cB的分子构成十分复杂,主要包括参与mRNA剪接所需、称为snRNP的核酸与蛋白质复合体的各种组分,以及snoRNAs、scaRNAs和其他相关蛋白质。此外,一些基因活化中不可或缺的转录因子、mRNA修饰的必需蛋白因子以及与细胞增殖和细胞周期功能相关的功能分子等,也被发现存在于CB之中或周边。其中,snRNAs、snoRNAs与scaRNAs等低分子量RNAs在CB中完成自身的成熟并且参与RNP的组装。染色体端粒长度的维持以及组蛋白基因簇的表观遗传修饰依赖于cB的形成与功能发挥。由此可见,CB与细胞周期、细胞增殖、应激和老化等一系列重要的细胞生物学过程存在密切的联系,同时其功能和表现的异常往往也提示与肿瘤的发生相关。该文在归纳cB相关研究最新进展的同时,着重讨论CB的结构与功能及其与疾病的潜在联系。     

甲藻染色体在非细胞体系核重建过程中核小体的组装

利用纯化的原始真核生物寇氏隐甲藻（Crypthecodinium cohnii E.）染色体,使之与非洲爪蟾（Xenopus laevis L.）S期卵提取物温育,发现甲藻染色体经历了一系列去凝集、再凝集的形态变化,最后形成类似典型高等真核生物的间期核结构。小球菌核酸酶酶切分析表明,不具备组蛋白和核小体结构的甲藻染色体在非洲爪蟾卵提取物中进行了核小体装配,此过程与DNA序列本身、核膜以及核纤层蛋白（Lamin）是否存在无关,但部分拓扑异构酶Ⅱ（TopoⅡ）参与了这个过程,说明核小体的组装并非为核重建所必需,决定染色体高级结构的因素并不在DNA本身,而可能是非细胞体系中的组蛋白和非组蛋白。     

体外装配核小体过程组蛋白浓度依赖的动力学模型

为探索组蛋白浓度对核小体体外装配的影响,本研究表达纯化了4种组蛋白,通过控制实验反应体系中组蛋白的浓度,利用盐透析法在体外装配了核小体,检测分析了组蛋白浓度与核小体组装效率的关系。以此实验数据为基础,提出了核小体组装过程组蛋白浓度依赖性的动力学模型。实验结果发现,反应体系中组蛋白浓度与核小体生成量呈典型的线性关系。依据动力学理论模型,进行线性回归拟合,回归系数达到0.963;经计算601 DNA序列组装核小体的反应速率常数k为1.49×10^-5mL·h·μg^-1。CS1序列验证动力学模型的线性回归相关系数为0.989,反应速率常数为1.52×10^-5mL·h·μg^-1。该实验方法及动力学模型中反应速率常数k可用于评价相同长度的DNA序列组装核小体的能力、组蛋白与其突变体以及组蛋白变体之间形成核小体结构能力的差异。该动力学模型的建立为理解核小体装配、核小体定位、染色质结构等相关问题提供了理论指导。     

着丝粒核小体结构研究进展

着丝粒是构成真核生物染色体的必需元件。在细胞有丝分裂或减数分裂时,微管通过动粒与染色体着丝粒连接,参与细胞分裂的染色体分离与分配过程,使染色体平均分配到子细胞中。构成着丝粒的基本单位是着丝粒特异的核小体,与常规核小体不同的是着丝粒核小体中的组蛋白H3被其变种——着丝粒组蛋白H3所替换。最近几年,着丝粒核小体的结构成为细胞生物学研究的热点之一。该文综述了最近在多种真核生物研究中,通过体外和体内实验,提出的着丝粒核小体结构的八聚体、六聚体、同型四聚体以及半八聚体模型,并对着丝粒核小体结构的动态模型与功能的关系进行了探讨。     

核小体定位的转录调控功能研究进展

核小体是真核生物染色质的基本组成单位,组蛋白八聚体在DNA 双螺旋上精确位置称为核小体定位．核小体定位已被证实在基因转录调控、DNA复制与修复、调控进化等过程中扮演着重要的角色．随着染色质免疫共沉淀-芯片(ChIP-chip)与染色质免疫共沉淀-测序(ChIP-seq)等高通量技术的出现,已测定了多种模式生物全基因组核小体定位图谱,掀起了一股核小体定位及其功能的研究热潮,并取得了一定的成果．本文介绍了核小体定位的概念,总结了核小体在启动子与编码区域内定位的基本模式．在此基础上,综述了核小体定位在转录起始、转录延伸、基因表达模式多样化以及可变剪接等方面的功能研究进展．     

核小体定位研究进展

核小体定位在诸如转录调控、DNA复制和修复等多种细胞过程中起着重要作用。基因组上核小体位置的确定涉及DNA、转录因子、组蛋白修饰酶和染色质重塑复合体之间的相互作用。核小体定位、组蛋白修饰、染色质重塑等问题已成为目前遗传学研究的热点——表观遗传学——的重要研究内容。文章从核小体定位基本概念、核小体定位与基因表达调控的关系、核小体定位实验研究和理论预测工作等几个方面总结了核小体定位的最新研究进展。     

两种模式生物核小体定位比较研究

在对两种模式生物酵母与果蝇胚胎期核小体定位进行研究时,发现不同物种间以及同一物种中不同表达模式基因上的核小体分布呈现出差显著异性。在总体上,转录起始位点附近的酵母核小体NFR区域比果蝇的NFR短。经基因中心对齐后,酵母与果蝇胚胎期沉默型基因的核小体缺失区域的两个边界中间处共同呈现了一个明确有着均匀间隔的核小体数n,且随着基因长度L的变长其周期性特性逐渐变模糊,但果蝇的图谱表现的更为复杂。结果表明,从单细胞酵母生物到多细胞果蝇生物间基因组的进化过程中,核小体组织的演化既有变异性,也具有保守性。     

核内不均一核糖核蛋白A2B1功能及其在病毒感染中的作用研究进展

核内不均一核糖核蛋白A2B1(hnRNPA2B1)是核不均一核糖核蛋白家族（hnRNPs）成员,在各种组织中广泛表达,具有识别和结合RNA和DNA模体的功能,其在细胞生命活动,如新生转录物的包装、选择性剪接和翻译调控中起重要作用。研究发现,hnRNPA2B1在病毒感染和抗病毒免疫方面扮演重要的角色,具有识别胞内病毒核酸,激发抗病毒免疫通路,调控病毒感染、复制及病毒释放等功能。本文就hnRNPA2B1的结构、功能,尤其是在DNA和RNA病毒感染中所涉及的作用机制作一综述。     

核小体定位与RNA剪接

根据核小体定位序列和缺失序列的碱基分布特征,应用多样性增量二次判别方法(IDQD)构建模型对这两类序列进行了区分,受试者操作特性曲线下的面积达到了0.958．应用这一模型研究了核小体在人类基因组剪接位点(GT/AG)邻近序列中的分布方式,发现外显子所对应的DNA序列通常倾向参与核小体的形成,并且由它所转录的RNA统计上具有较强的刚性,而剪接位点及其邻近的内含子对应的DNA序列则避免参与核小体的形成,所转录的RNA统计上具有较强的柔性．进一步还发现,DNA序列的核小体定位/缺失和RNA的刚性/柔性具有统计相关性,为从机制上解释为何前体RNA剪接事件与DNA序列中的核小体定位信息有关提供了依据．     

基于DNA变形能的核小体定位预测方法研究进展

真核细胞中,作为染色质基本结构单元的核小体参与调控基因的转录、DNA复制、重组以及RNA剪接等诸多生物学过程。阐明核小体定位机制并准确预测核小体在染色体上的位置对解读染色质结构与功能有重要生物学意义。在过去30多年时间里,研究人员发展了多种预测核小体位置的方法。最理想的方法应考虑DNA序列、组蛋白修饰和染色质重塑等影响核小体定位的诸多因素,然而现实中,捕捉主要因素的模型也往往具有很高的鲁棒性和实用价值。DNA序列偏好性是在全基因组尺度上影响核小体定位的最重要因素之一,因此基于DNA序列的核小体定位预测方法也最常见。这种方法可大致分为两类,即基于DNA序列信息的生物信息学模型和基于DNA变形能的生物物理学模型。本文重点介绍生物物理学模型近些年取得的主要进展。     

Composition of nuclear dense bodies and nucleolus-associated bodies in interphase nuclei of the unicellular green alga Chlamydomonas reinhardtii

Summary— The interphase nucleus of the green alga, Chlamydomonas reinhardtii, displayed two types of bodies some of them, the dense bodies, lying apparently free in the nucleoplasm while the others were attached to the nucleolus and were, therefore, referred to as nucleolus-associated bodies (NABs). The presence of DNA, RNA and histones in dense bodies was investigated by means of post-embedding immunocytochemistry and cytochemistry using a monoclonal antibody to single and double stranded DNA, a polyclonal antibody to rye H3 histones and RNase A-gold complexes. The dense bodies were shown to contain significant amounts of RNA but neither DNA nor histones were detected; their composition was thus similar to that of the dense bodies described in higher plant cells. We propose that dense bodies might be implicated in the assembly of the 25 to 45 nm granules observed throughout the nucleoplasm of Chalamydomonas interphase nuclei. The composition of NABs was found to be distinct from that of the dense bodies since they were labeled by the antibody to DNA, specially in cryofixed and cryosubstituted specimens. The presence of DNA in NABs together with their intimate association to the nucleolus suggest that they may correspond to specific segments of chromosomes.     

A role for nuclear lamins in nuclear envelope assembly.

The molecular interactions responsible for nuclear envelope assembly after mitosis are not well understood. In this study, we demonstrate that a peptide consisting of the COOH-terminal domain of Xenopus lamin B3 (LB3T) prevents nuclear envelope assembly in Xenopus interphase extracts. Specifically, LB3T inhibits chromatin decondensation and blocks the formation of both the nuclear lamina-pore complex and nuclear membranes. Under these conditions, some vesicles bind to the peripheral regions of the chromatin. These "nonfusogenic" vesicles lack lamin B3 (LB3) and do not bind LB3T; however, "fusogenic" vesicles containing LB3 can bind LB3T, which blocks their association with chromatin and, subsequently, nuclear membrane assembly. LB3T also binds to chromatin in the absence of interphase extract, but only in the presence of purified LB3. Additionally, we show that LB3T inhibits normal lamin polymerization in vitro. These findings suggest that lamin polymerization is required for both chromatin decondensation and the binding of nuclear membrane precursors during the early stages of normal nuclear envelope assembly.     

Relation between nuclear envelope and nuclear lamina in nuclear assembly in vitro

Xenopus laevis egg extracts cell-free nuclear assembly system was used as an experimental model to study the process of nuclear lamina assembly in nuclear reconstitution in vitro. The experimental results showed that lamin was involved in the nuclear assembly in vitro. The assembly of nuclear lamina was preceded by the assembly of nuclear matrix, and probably, inner nuclear matrix assembly provided the basis for nuclear lamina assembly. Inhibition of normal assembly of nuclear lamina, by preincubating egg extracts cell-free system with anti-lamin antibodies, resulted in abnormal assembly of nuclear envelope, suggesting that nuclear envelope assembly is closely associated with nuclear lamina assembly.     

Nuclear development in locust fat body: the influence of juvenile hormone on inclusion bodies and the nuclear matrix

The hormonal induction of vitellogenesis in insects and in oviparous vertebrates are prime models of gene regulation in eukaryotes. In vertebrates the process is under estrogenic control and normally confined to females, although males can be artificially induced. In locust in contrast, juvenile hormone (JH) is central to fat body development in both males and females, yet the response is strongly sex limited not only for vitellogenin production but also in terms of total protein, DNA and RNA synthesis and nuclear ploidy levels. To differentiate further possible sex and/or JH related developmental aspects in locusts, large-scale nuclear events were examined during normal adult maturation and in animals treated with antiallatropins and JH analogs. Fat body nuclei undergo extensive restructuring during normal development in both sexes. This included progressive nuclear enlargement, accompanied by extensive proliferation of nuclear matrix components and elaboration of complex inclusion bodies (NB). The isolated protein matrix was unusually complex relative to similar structures from vertebrates and the NB were firmly anchored to it. Although matrix proteins were qualitatively similar to those from other sources, as assessed by SDS polyacrylamide gel electrophoresis, several major matrix polypeptides, including lamins A and B, and components greater than 150 kD, fluctuated quantitatively during development and in concert with nuclear enlargement. The number and morphology of the NB were unrelated to sex, but increased in direct proportion to absolute nuclear volumes. All changes were more pronounced in females, where higher ploidy levels, larger nuclei and correspondingly more internal matrix elements occurred. Suppression of JH production by precocene prevented all foregoing nuclear changes, but re-exposure to methoprene rapidly induced normal development. The results are compared to analogous nuclear changes in steroid responsive vertebrate tissues.