New advances of microRNAs in the pathogenesis of rheumatoid arthritis,with a focus on the crosstalk between DNA methylation and the microRNA machinery

Rheumatoid arthritis (RA) is a symmetrical polyarticular disease of unknown aetiology that affects primarily the articular cartilage and bone. Characteristic features of RA pathogenesis are persistent inflammation, synovium hyperplasia and cartilage erosion accompanied by joint swelling and joint destruction. Several lines of evidence have showed a crucial role of activated fibroblast-like synoviocytes (FLS) in the pathogenesis of RA. MicroRNAs (miRNAs) are endogenous, single-stranded, non-coding RNAs with about 21 nucleotides in length and have been detected in a variety of sources, including tissues, serum, and other body fluids, such as saliva. In light of key roles of miRNAs in the regulation of gene expression, miRNAs influence a wide range of physiological and pathological processes. For example, miRNAs are evident in various malignant and nonmalignant diseases, and accumulating evidence also shows that miRNAs have important roles in the pathogenesis of RA. It has been demonstrated that miRNAs can be aberrantly expressed even in the different stages of RA progression, allowing miRNAs to help understand the pathogenesis of the disease, to act as important biomarkers, and to monitor the disease severity and the effects of drug treatment. In addition, miRNAs are emerging as potential targets for new therapeutic strategies of this kind of autoimmune disorders. The ultimate goal is the identification of miRNA targets that could be manipulated through specific therapies, aiming at activation or inhibition of specific miRNAs responsible for the RA development. In this review, the importance of miRNAs in the pathogenesis of RA is discussed systematically, with particular emphasis on the role of the crosstalk between DNA methylation and the microRNA machinery.     

Expression profile analysis of microRNAs during hair follicle development in the sheep foetus

MicroRNAs (miRNAs) regulate the development and growth cycle of hair follicles (HFs). The molecular mechanism by which miRNAs determine the development of HFs in the sheep foetus remains elusive. In this study, the expression profiles of miRNAs at 11 development periods (45, 55, 65, 75, 85, 95, 105, 115, 125, 135 and 145 d) in sheep foetus skin were analysed by high-throughput sequencing and bioinformatics analysis. A total of 72 conserved miRNAs, 44 novel miRNAs and 32 known miRNAs were significantly differentially expressed. qRT-PCR results for 18 miRNAs were consistent with the sequencing data. 85 d of foetal development was the starting point for secondary hair follicle (SF) development according to tissue morphology and cluster analysis. In SF development, the prolactin signalling pathway and platelet activation played important roles, and 10 miRNAs were potential candidate miRNAs in SF initiation.     

Identification and differential expression of microRNAs in the ovaries of pigs (Sus scrofa) with high and low litter sizes

Litter size affects profitability in the swine industry. Mammalian ovaries play important roles during reproduction, including ovulation and hormone secretion, which are tightly regulated by specific microRNAs (miRNAs). In this study, we investigated the effects of specific miRNAs on porcine litter size. We compared the ovarian miRNAs of Yorkshire pigs with high (YH) and low (YL) litter sizes using Solexa sequencing technology. We identified 327 and 320 miRNAs in the ovaries of YH and YL pigs respectively. A total of 297 miRNAs were co‐expressed; 30 and 23 miRNAs respectively were specifically expressed in the two libraries. A total of 83 novel miRNAs were predicted; 37 specific miRNAs were obtained, of which 21 miRNAs were upregulated and 16 miRNAs were downregulated in YH compared with YL. Additionally, 19 628 and 19 250 target genes were predicted in the two libraries respectively. The results revealed that specific miRNAs (i.e., miR‐224, miR‐99a, let‐7c, miR‐181c, miR‐214 and miR‐21) may affect porcine litter size. The results of this study will help in gaining understanding of the role of miRNAs in porcine litter size regulation.     

微小RNA与细胞凋亡的研究进展

微小RNA(miRNAs)是最近发现的由18~24个核苷酸组成的RNA,通过对目标mRNA的抑制而发挥重要的调节作用。目前所有已研究的多细胞真核生物表明它们是通过miRNAs来调节细胞基本的生理功能,这些功能包括细胞的增殖、分化和死亡。本文讨论了miRNAs在调节细胞增殖和凋亡方面的功能:其中,抗凋亡的miRNAs有miR-17家族、miR-21、bantam和miR-14;促凋亡的miRNAs有let-7、miR-15a和miR-16。     

微小RNAs在肝癌中的作用机制及相关应用研究进展

微小RNAs(miRNAs)是一类内源性小型非编码RNA,可通过调控靶基因表达参与大多数生物学过程。近年来,miRNAs在肝癌发生发展进程中相关作用机制的研究逐渐深入,miRNAs作为其中关键调控因子和主要参与者,已成为肝癌早期诊断、靶向治疗和预后评估中的一个关键靶标。本文着重强调miRNAs在肝癌发生发展、多重耐药性中的作用以及作为肝癌潜在治疗靶点的价值,并就miRNAs在肝癌中的功能、分子作用通路以及应用三方面的相关研究进展进行综述。     

Signature miRNAs involved in the innate immunity of invertebrates

The innate immune system, including the cell-based immunity (mainly apoptosis and phagocytosis) and the humoral immunity (such as pro-phenoloxidase system), is the first defense line of animals against the infection of pathogens in a non-specific manner, which is fine regulated through the gene expression regulations. The microRNAs (miRNAs) are recognized as important regulators of gene expression. To date, however, a comprehensive view about the regulation of innate immunity by miRNAs is not available. To address this issue, the signature miRNAs involved in the innate immunity were characterized in this study. The phagocytosis, apoptosis and phenoloxidase (PO), a key enzyme in the pro-phenoloxidase system, of invertebrate shrimp were activated or inhibited, followed by the small RNA sequencing. The results showed that a total of 24 miRNAs took great effects on phagocytosis, apoptosis or the pro-phenoloxidase system, which were further confirmed by Northern blots. Among the 24 innate immunity-associated miRNAs, 21 miRNAs were conserved in animals, suggesting that these miRNAs might share the similar or the same functions in different species of animals. Based on degradome sequencing and prediction of target genes, it was found that the miRNAs might mediate the regulations of phagocytosis, apoptosis or pro-phenoloxidase system by targeting different genes. Therefore our study presented the first comprehensive view of the miRNAs associated with innate immunity, which would facilitate to reveal the molecular events in the regulation of innate immunity.     

Identification and validation of Asteraceae miRNAs by the expressed sequence tag analysis

MicroRNAs (miRNAs) are small non-coding RNA molecules that play a vital role in the regulation of gene expression. Despite their identification in hundreds of plant species, few miRNAs have been identified in the Asteraceae, a large family that comprises approximately one tenth of all flowering plants. In this study, we used the expressed sequence tag (EST) analysis to identify potential conserved miRNAs and their putative target genes in the Asteraceae. We applied quantitative Real-Time PCR (qRT-PCR) to confirm the expression of eight potential miRNAs in Carthamus tinctorius and Helianthus annuus. We also performed qRT-PCR analysis to investigate the differential expression pattern of five newly identified miRNAs during five different cotyledon growth stages in safflower. Using these methods, we successfully identified and characterized 151 potentially conserved miRNAs, belonging to 26 miRNA families, in 11 genus of Asteraceae. EST analysis predicted that the newly identified conserved Asteraceae miRNAs target 130 total protein-coding ESTs in sunflower and safflower, as well as 433 additional target genes in other plant species. We experimentally confirmed the existence of seven predicted miRNAs, (miR156, miR159, miR160, miR162, miR166, miR396, and miR398) in safflower and sunflower seedlings. We also observed that five out of eight miRNAs are differentially expressed during cotyledon development. Our results indicate that miRNAs may be involved in the regulation of gene expression during seed germination and the formation of the cotyledons in the Asteraceae. The findings of this study might ultimately help in the understanding of miRNA-mediated gene regulation in important crop species.     

Discrete clusters of virus-encoded micrornas are associated with complementary strands of the genome and the 7.2-kilobase stable intron in murine cytomegalovirus

The prevalence and importance of microRNAs (miRNAs) in viral infection are increasingly relevant. Eleven miRNAs were previously identified in human cytomegalovirus (HCMV); however, miRNA content in murine CMV (MCMV), which serves as an important in vivo model for CMV infection, has not previously been examined. We have cloned and characterized 17 novel miRNAs that originate from at least 12 precursor miRNAs in MCMV and are not homologous to HCMV miRNAs. In parallel, we applied a computational analysis, using a support vector machine approach, to identify potential precursor miRNAs in MCMV. Four of the top 10 predicted precursor sequences were cloned in this study, and the combination of computational and cloning analysis demonstrates that MCMV has the capacity to encode miRNAs clustered throughout the genome. On the basis of drug sensitivity experiments for resolving the kinetic class of expression, we show that the MCMV miRNAs are both early and late gene products. Notably, the MCMV miRNAs occur on complementary strands of the genome in specific regions, a feature which has not previously been observed for viral miRNAs. One cluster of miRNAs occurs in close proximity to the 5' splice site of the previously identified 7.2-kb stable intron, implying a variety of potential regulatory mechanisms for MCMV miRNAs.     

Identification and comparative analysis of the ovary and testis microRNAome of mud crab Scylla paramamosain

The role of microRNA (miRNA) in reproductive regulation is attracting increasingly more attention. In this study, we obtained 9,643,114 and 15,498,999 raw reads from the ovary and testis library of important farmed mud crab Scylla paramamosain, respectively. After data mining, a total of 4,096,464 and 11,737,973 mappable small RNA sequences remained for analysis. By mapping to the reference genome and expressed sequence tag (EST) of Daphnia pulex and other crabs, a total of 1,417 miRNAs were identified. On the basis of 1,417 miRNAs, 514 (36.3%) unique miRNAs coexpressed in the gonad of female and male libraries, and 336 (23.7%) and 567 (40%) expressed preferentially in female and male libraries, respectively. Analysis of library sequencing data resulted in the identi?cation of 108 miRNAs (out of 1,417; 7.6%) that showed signi?cant differential expression between the two samples. Of these, 13 miRNAs were expressed only in the testis, two miRNAs were expressed only in the ovary, and 93 miRNAs were coexpressed: 57 (61.3%) were upregulated (ovary/testis) and 36 (38.7%) were downregulated (ovary/testis). To confirm the expression patterns of the predicted miRNAs, we randomly selected 14 candidate miRNAs from 108 differentially expressed miRNAs and performed stem–loop real time quantitative PCR (RT‐qPCR) assays in five ovary developing stages. Five miRNAs showed similar expression patterns in almost every stage as those revealed by identification of differentially expressed genes (IDEG6) analysis. The above five miRNAs were predicted to match the 3′‐untranslated region of the published S. paramamosain gene. Four out of five miRNA had a regulation effect on many genes, especially the genes related to gonadal development.     

Identification and Expression Analysis of microRNAs at the Grain Filling Stage in Rice(Oryza sativa L.)via Deep Sequencing

MicroRNAs (miRNAs) have been shown to play crucial roles in the regulation of plant development. In this study, high-throughput RNA-sequencing technology was used to identify novel miRNAs, and to reveal miRNAs expression patterns at different developmental stages during rice (Oryza sativa L.) grain filling. A total of 434 known miRNAs (380, 402, 390 and 392 at 5, 7, 12 and 17 days after fertilization, respectively.) were obtained from rice grain. The expression profiles of these identified miRNAs were analyzed and the results showed that 161 known miRNAs were differentially expressed during grain development, a high proportion of which were up-regulated from 5 to 7 days after fertilization. In addition, sixty novel miRNAs were identified, and five of these were further validated experimentally. Additional analysis showed that the predicted targets of the differentially expressed miRNAs may participate in signal transduction, carbohydrate and nitrogen metabolism, the response to stimuli and epigenetic regulation. In this study, differences were revealed in the composition and expression profiles of miRNAs among individual developmental stages during the rice grain filling process, and miRNA editing events were also observed, analyzed and validated during this process. The results provide novel insight into the dynamic profiles of miRNAs in developing rice grain and contribute to the understanding of the regulatory roles of miRNAs in grain filling.     

Identification of microRNAs of the herpesvirus family

Epstein-Barr virus (EBV or HHV4), a member of the human herpesvirus (HHV) family, has recently been shown to encode microRNAs (miRNAs). In contrast to most eukaryotic miRNAs, these viral miRNAs do not have close homologs in other viral genomes or in the genome of the human host. To identify other miRNA genes in pathogenic viruses, we combined a new miRNA gene prediction method with small-RNA cloning from several virus-infected cell types. We cloned ten miRNAs in the Kaposi sarcoma-associated virus (KSHV or HHV8), nine miRNAs in the mouse gammaherpesvirus 68 (MHV68) and nine miRNAs in the human cytomegalovirus (HCMV or HHV5). These miRNA genes are expressed individually or in clusters from either polymerase (pol) II or pol III promoters, and share no substantial sequence homology with one another or with the known human miRNAs. Generally, we predicted miRNAs in several large DNA viruses, and we could neither predict nor experimentally identify miRNAs in the genomes of small RNA viruses or retroviruses.     

Combined Small RNA and Degradome Sequencing Reveals Novel MiRNAs and Their Targets in the High-Yield Mutant Wheat Strain Yunong 3114

Wheat is one of the main food sources worldwide; large amount studies have been conducted to improve wheat production. MicroRNAs (miRNAs) with about 20–30 nucleotide are a class of regulatory small RNAs (sRNAs), which could regulate gene expression through sequence-specific base pairing with target mRNAs, playing important roles in plant growth. An ideal plant architecture (IPA) is crucial to enhance yield in bread wheat. In this study, the high-yield wheat strain Yunong 3114 was EMS-mutagenesis from the wild-type strain Yunong 201, exhibiting a preferable plant structure compared with the wild-type strain. We constructed small RNA and degradome libraries from Yunong 201 and Yunong 3114, and performed small RNA sequencing of these libraries in order identify miRNAs and their targets related to IPA in wheat. Totally, we identified 488 known and 837 novel miRNAs from Yunong 3114 and 391 known and 533 novel miRNAs from Yunong 201. The number of miRNAs in the mutant increased. A total of 37 known and 432 putative novel miRNAs were specifically expressed in the mutant strain; furthermore, 23 known and 159 putative novel miRNAs were specifically expressed in the wild-type strain. A total of 150 known and 100 novel miRNAs were differentially expressed between mutant and wild-type strains. Among these differentially expressed novel miRNAs, 4 and 8 predict novel miRNAs were evidenced by degradome sequencing and showed up-regulated and down-regulated expressions in the mutant strain Yunong 3114, respectively. Targeted gene annotation and previous results indicated that this set of miRNAs is related to plant structure. Our results further suggested that miRNAs may be necessary to obtain an optimal wheat structure.     

乳腺发育及泌乳相关miRNA研究进展

MicroRNA(miRNA)是一种重要的转录后调控的非编码RNA, 参与调控哺乳动物乳腺发育和泌乳。文章总结了乳腺发育和泌乳过程中miRNA表达的时空特异性, 综述了个别miRNA对乳腺发育和泌乳的调节作用, 旨在为乳腺miRNA的研究提供指导, 为利用miRNA促进乳腺健康发育和调控高质高效产乳提供理论基础和研究思路。     

MicroRNA expression in the aging mouse thymus

MicroRNAs (miRNAs) have been implicated in the process of aging in many model organisms, such as Caenorhabditis elegans, and in many organs, such as the mouse lung and human epididymis. However, the role of miRNAs in the thymus tissues of the aging mouse remains unclear. To address this question, we investigated the miRNA expression profiles in the thymuses of 1-, 10- and 19-month-old mice using miRNA array and qRT-PCR assays. A total of 223 mouse miRNAs were screened, and the expression levels of those miRNAs exhibited gradual increases and decreases over the course of thymus aging. Fifty miRNAs in the 10-month-old thymus and 81 miRNAs in the 19-month-old thymus were defined as differentially expressed miRNAs (p < 0.05) in comparison with their levels in the 1-month-old mouse, and approximately one-third of these miRNAs were grouped within 11 miRNA clusters. Each miRNA cluster contained 2 to 5 miRNA genes, and most of the cluster members displayed similar expression patterns, being either increased or decreased. In addition, Ingenuity Pathway Analysis (IPA) software and the IPA database were used to analyze the 12 miRNAs that exhibited significant expression changes, revealing that as many as 15 pathways may be involved. Thus, our current study determined the expression profiles of miRNAs in the mouse thymus during the process of aging. The results suggested that these miRNAs could become meaningful biomarkers for studying thymus aging and that the aging-related alternations in miRNA expression may be involved in the regulation of cell proliferation, apoptosis, development and carcinogenesis/tumorigenesis.