小RNAs在生殖细胞发育过程中的作用

很多动物可以产生具调节作用的小RNAs,根据产生方式和作用机制可以将它们分为三类:微小RNAs(miRNAs)、与Piwi相互作用的RNAs(piRNAs)和内源小干扰RNAs(endo-siRNAs),这些小RNAs可以在生物生殖细胞发育过程中发挥重要作用。其中miRNAs的主要作用是调节蛋白质基因的表达;piRNAs主要的作用是沉默转座因子,但piRNAs主要存在于生殖细胞中;endo-siRNAs则可能具有上述两种主要作用。该文论述了这三种小RNAs在生物生殖细胞发育过程中的作用,同时也讨论了它们在治疗生物不育及其在生物节育方面的应用前景。     

植物小RNAs的研究进展

小RNAs(长度小于40 nt)是nc-RNAs重要的一部分,现在植物中已发现了多种小RNAs,如小干扰RNAs(siRNAs)、微小RNAs(miRNAs)、反式作用的小干扰RNAs、天然反义转录小干扰RNAs、异染色质小干扰RNAs、长小片段小干扰RNAs、天然反义转录的微小RNAs及其一些未命名的小RNAs.成熟的小RNAs聚集相关的蛋白质因子,可以抑制转录,导致转录水平的基因沉默(TGS);或介导目标mRNA的剪切,抑制翻译,导致转录后水平基因沉默(PTGS).就这些植物小RNAs产生及其作用的研究进展作一概述     

天然反义转录物及其调控基因的表达机制

天然反义转录(NATs)是一组编码蛋白质或非编码蛋白质的RNAs, 与其他(有义)转录物具有互补序列, 可以调节有义链的表达。这种调节可以发生在转录水平或转录后水平, 调节方式有转录干扰、RNA封闭、双链依赖机制或染色质重建(修饰)等。正义链和反义链分别加工成小RNAs调节基因表达, 也是NATs调节基因表达的重要方式, 如piRNAs的“乒乓机制”。实验或计算机研究已经证明了NATs在生物中广泛存在, 是一种重要的基因表达调节方式。文章论述了NATs的重要作用和机理, 重点论述了NATs的调节机制和相关的小RNAs。     

小RNA的生成机制与功能

小RNA长度在20~32 nt之间,通过染色质修饰、mRNA降解和翻译抑制来调控基因表达。小RNA可以分为三类:小干扰RNA、微小RNA和piRNAs。小干扰RNA主要抵御转座子和病毒的侵袭。微小RNA的表达受发育水平调控且有组织特异性,在发育和细胞分化中起作用。piRNAs在生殖细胞和干细胞中表达,可使反转座子沉默。综述了这几种小RNA的定义与分类、生成机制、功能及其研究方法。     

小RNAs作用机制的研究进展

RNAi的发现引发了生物学的一次革命, 也揭示了一种原来未被发现的, 通过小RNAs(大小~20–30 nt)家族在转录水平或转录后水平调解基因表达的方式。在真核生物中, 这些小RNAs包括siRNAs、miRNAs、piRNAs、scnRNAs、21U-RNAs和其他一些小RNAs等。它们通过调节基因表达来控制细胞的代谢、生长和分化, 维持基因组的完整性, 协调生殖细胞的成熟和抑制病毒对细胞的侵袭以及转座成分的转座。文章综述了这些小RNAs在鉴定和生物合成方面的研究进展, 并讨论了它们对基因表达的调节作用。     

piRNA/PIWI功能调控与精子发生

piRNAs(PIWI-interacting RNAs)是一类与PIWI相互作用的小非编码RNAs(small noncoding RNAs, sncRNAs),其长度介于24~32 nt,特异性地在动物生殖腺细胞中表达。近来研究表明piRNA/PIWI系统在动物生殖腺细胞的基因组转座元件沉默及转录后调控mRNAs方面具有重要功能。最近,中国科学院上海生物化学与细胞生物学研究所刘默芳课题组的一项研究表明,在人和小鼠的精子发生过程中,PIWI (鼠源同源蛋白MIWI、人源同源蛋白HIWI)的严格代谢调控至关重要。以此为契机,本文综述了piRNA/PIWI在哺乳动物(主要是小鼠和人)精子发生过程中调控功能的研究进展。     

RNA沉默在植物生物逆境反应中的作用

RNA沉默是真核生物共有的基因表达调节机制和防御机制。在植物RNA沉默中, 一些小RNAs, 如微小 RNAs和小干扰RNAs, 在植物防御病毒、细菌或食草动物的反应中具有重要作用。为了抑制宿主的RNA沉默系统, 植物病毒或细菌进化出了在RNA沉默不同阶段起作用的病毒沉默抑制子或细菌沉默抑制子, 来克服寄主的RNA沉默反应。文章就植物RNA沉默、病毒沉默抑制子、细菌沉默抑制子及其相关防御反应的一些新进展做一概述。     

小RNAs在沉默转座因子中的作用

在很多生物基因组中都存在DNA成分的转座序列,它们能够转座到基因组的很多位点,对基因组造成很大的危害,如破坏编码基因、改变基因表达的调节网络、使染色体断裂或造成大范围基因重排等。真核生物已经进化出了多种机制来控制这些寄生核酸序列造成的损伤,以维持基因组完整性。虽然这些机制在不同生物中有些差异,但其中一种主要的机制是通过小RNAs介导的,这些小RNAs包括小干扰RNAs、piwi相互作用的小RNAs、微小RNAs、扫描RNAs和21U-RNAs等。这些小RNAs可以通过DNA水平剪切转座序列,或在转录和(或)转录后水平沉默转座成分。该文就这些小RNAs沉默转座成分的机制和功能做一论述。     

家蚕微孢子虫中小RNAs的全基因组鉴定与分析

【目的】家蚕Bombyx mori微粒子病是蚕业生产上的毁灭性病害,家蚕微孢子虫Nosema bombycis是该病的病原,可经卵垂直传播和经口水平传播。为了探索家蚕微孢子虫中对重复元件的抵御以及对基因转录调控的潜在方式,本研究拟在基因组水平上对该物种的小RNAs进行全面系统的分析,鉴定与转座子相关的小RNAs和潜在的miRNAs。【方法】从感染家蚕微孢子虫的家蚕中肠中提取总RNA,分离小片段RNA并反转录后,进行Solexa高通量测序。通过生物信息学方法对小RNAs进行分类及功能注释,鉴定起源于家蚕微孢子虫不同类型转座子的小RNAs,并对潜在的miRNA进行预测分析。【结果】家蚕微孢子虫小RNAs的长度主要是24和25 nt,其中大部分序列表现出5′末端的尿嘧啶偏好性。家蚕微孢子虫中存在丰富的与转座子相关联的小RNAs,并且与转座子标准序列匹配的反义小RNAs明显多于正义小RNAs。同时,鉴定获得了31个候选miRNAs,部分为Nosema属的其他孢子虫中所共有,暗示其在微孢子虫基因组进化上具有保守性。【结论】首次鉴定到家蚕微孢子虫的转座子相关性小RNAs,暗示小RNAs在家蚕微孢子虫基因组对转座子防御过程中起到作用,31个潜在的miRNAs为家蚕微孢子虫miRNAs的功能验证提供了后续靶标。     

家蚕微孢子虫中小RNAs的全基因组鉴定与分析（英文）

【目的】家蚕Bombyx mori微粒子病是蚕业生产上的毁灭性病害,家蚕微孢子虫Nosema bombycis是该病的病原,可经卵垂直传播和经口水平传播。为了探索家蚕微孢子虫中对重复元件的抵御以及对基因转录调控的潜在方式,本研究拟在基因组水平上对该物种的小RNAs进行全面系统的分析,鉴定与转座子相关的小RNAs和潜在的miRNAs。【方法】从感染家蚕微孢子虫的家蚕中肠中提取总RNA,分离小片段RNA并反转录后,进行Solexa高通量测序。通过生物信息学方法对小RNAs进行分类及功能注释,鉴定起源于家蚕微孢子虫不同类型转座子的小RNAs,并对潜在的miRNA进行预测分析。【结果】家蚕微孢子虫小RNAs的长度主要是24和25 nt,其中大部分序列表现出5'末端的尿嘧啶偏好性。家蚕微孢子虫中存在丰富的与转座子相关联的小RNAs,并且与转座子标准序列匹配的反义小RNAs明显多于正义小RNAs。同时,鉴定获得了31个候选miRNAs,部分为Nosema属的其他孢子虫中所共有,暗示其在微孢子虫基因组进化上具有保守性。【结论】首次鉴定到家蚕微孢子虫的转座子相关性小RNAs,暗示小RNAs在家蚕微孢子虫基因组对转座子防御过程中起到作用,31个潜在的miRNAs为家蚕微孢子虫miRNAs的功能验证提供了后续靶标。     

Transposon Defense by Endo-siRNAs,piRNAs and Somatic pilRNAs in Drosophila: Contributions of Loqs-PD and R2D2

Transposable elements are a serious threat for genome integrity and their control via small RNA mediated silencing pathways is an ancient strategy. The fruit fly Drosophila melanogaster has two silencing activities that target transposons: endogenous siRNAs (esiRNAs or endo-siRNAs) and Piwi-interacting small RNAs (piRNAs). The biogenesis of endo-siRNAs involves the Dicer-2 co-factors Loqs-PD, which acts predominantly during processing of dsRNA by Dcr-2, and R2D2, which primarily helps to direct siRNAs into the RNA interference effector Ago2. Nonetheless, loss of either protein is not sufficient to produce a phenotype comparable with a dcr-2 mutation. We provide further deep sequencing evidence supporting the notion that R2D2 and Loqs-PD have partially overlapping function. Certain transposons display a preference for either dsRBD-protein during production or loading; this appeared to correlate neither with overall abundance, classification of the transposon or a specific site of genomic origin. The endo-siRNA biogenesis pathway in germline operates according to the same principles as the existing model for the soma, and its impairment does not significantly affect piRNAs. Expanding the analysis, we confirmed the occurrence of somatic piRNA-like RNAs (pilRNAs) that show a ping-pong signature. We detected expression of the Piwi-family protein mRNAs only barely above background, indicating that the somatic pilRNAs may arise from a small sub-population of somatic cells that express a functional piRNA pathway.     

Mice deficient for a small cluster of Piwi-interacting RNAs implicate Piwi-interacting RNAs in transposon control

The mammalian testis expresses a class of small noncoding RNAs that interact with mammalian PIWI proteins. In mice, the PIWI-interacting RNAs (piRNAs) partner with mammalian PIWI proteins, PIWIL1 and PIWIL2, also known as MIWI and MILI, to maintain transposon silencing in the germline genome. Here, we demonstrate that inactivation of Nct1/2, two noncoding RNAs encoding piRNAs, leads to derepression of LINE-1 (L1) but does not affect mouse viability, spermatogenesis, testicular gene expression, or fertility. These findings indicate that piRNAs from a cluster on chromosome 2 are necessary to maintain transposon silencing.     

Bombyx small RNAs: genomic defense system against transposons in the silkworm, Bombyx mori

Selfish genetic elements called transposons can insert themselves at new locations in host genomes to modify gene structure and alter gene expression. Expansion of transposons can occur when novel transposition events are transmitted to subsequent generations after germline hopping. Therefore, organisms seem likely to have evolved defense mechanisms to silence transposons in the germline. Recently, small RNAs interacting with Piwi proteins (piwi-interacting RNAs: piRNAs) have been demonstrated to be involved in genomic defense mechanism against transposons. Here, we show that piRNA-like small RNAs are present abundantly in the Bombyx ovary. We cloned 38,493 kinds of Bombyx small RNA from the ovary and performed functional characterization. Bombyx small RNAs showed a unimodal length distribution with a peak at 28nt and a strong bias for U at the 5' end. We found that 12,869 kinds of Bombyx small RNAs were associated with transposons or repetitive sequences. We classified them as repeat-associated small interfering RNAs (rasiRNAs), a subclass of piRNAs. Notably, antisense rasiRNAs have a strong bias toward U at 5' ends; in contrast, sense rasiRNAs have a strong bias toward A at nucleotide position 10, indicating that the piRNA amplification loop proposed in Drosophila is evolutionarily conserved in Bombyx. These results suggest that Bombyx small RNAs regulate transposon activity.     

Mighty Piwis defend the germline against genome intruders

Piwis are a germline-specific subclass of the Argonaute family of RNA interference (RNAi) effector proteins that are associated with a recently discovered group of small RNAs (piRNAs). Recent studies in Drosophila and zebrafish directly implicate Piwi proteins in piRNA biogenesis to maintain transposon silencing in the germline genome (Brennecke et al., 2007; Gunawardane et al., 2007; Houwing et al., 2007). This function may be conserved in mice as loss of Miwi2, a mouse Piwi homolog, leads to germline stem cell and meiotic defects correlated with increased transposon activity (Carmell et al., 2007).     

The piRNA pathway: a fly's perspective on the guardian of the genome

Throughout the eukaryotic lineage, small RNA silencing pathways protect the genome against the deleterious influence of selfish genetic elements such as transposons. In animals an elaborate small RNA pathway centered on PIWI proteins and their interacting piRNAs silences transposons within the germline. In contrast to other small RNA silencing pathways, we lack a mechanistic understanding of this genome defense system. However, genetic and molecular studies have uncovered a fascinating conceptual framework for this pathway that is conserved from sponges to mammals. We discuss our current understanding of the piRNA pathway in Drosophila with an emphasis on origin and biogenesis of piRNAs.     

Adaptation to P element transposon invasion in Drosophila melanogaster

Transposons evolve rapidly and can mobilize and trigger genetic instability. Piwi-interacting RNAs (piRNAs) silence these genome pathogens, but it is unclear how the piRNA pathway adapts to invasion of new transposons. In Drosophila, piRNAs are encoded by heterochromatic clusters and maternally deposited in the embryo. Paternally inherited P element transposons thus escape silencing and trigger a hybrid sterility syndrome termed P-M hybrid dysgenesis. We show that P-M hybrid dysgenesis activates both P elements and resident transposons and disrupts the piRNA biogenesis machinery. As dysgenic hybrids age, however, fertility is restored, P elements are silenced, and P element piRNAs are produced de novo. In addition, the piRNA biogenesis machinery assembles, and resident elements are silenced. Significantly, resident transposons insert into piRNA clusters, and these new insertions are transmitted to progeny, produce novel piRNAs, and are associated with reduced transposition. P element invasion thus triggers heritable changes in genome structure that appear to enhance transposon silencing.