Identification and characterization of differentially expressed novel miRNAs (21–24 nt) in a Macrophomina phaseolina resistant RIL line of jute (Corchorus capsularis L.)

A RIL population of jute developed by crossing one resistant accession CIM 036 and a susceptible variety JRC 412 was used to identify novel defence related miRNAs activated upon challenged inoculation with stem rot pathogen Macrophomina phaseolina. About a total of 15.7 million reads were generated from the resistant line with the read length of 1 × 50 bp. Here, we identified nine mature novel microRNAs which passed Minimum Free Energy (MFE Kcal/mol) criteria. Target site and secondary structure were predicted and most of them showed ubiquitination and selective autophagy activity with high expression value. Five novel miRNAs viz. Candidate_41, Candidate_9, Candidate_66, Candidate_65 and Candidate_8 had free energy less than −25 kcal/mol. Known microRNAs viz. miR-845b and miR-166 superfamily are abundantly expressed with high expression value. The sequence of jute miR-845b superfamily is identical to that of Arabidopsis thaliana except at 18th position, but unlike in A. thaliana it targets the coding sequence for the P-loop motif in the mRNA sequences for disease resistance proteins with nucleotide binding site (NBS) and leucine-rich repeat (LRR) motifs. In-silico analysis suggested that miR-845b and miR-166 superfamily provided NBS-LRR and ROS mediated defence and subsequently expression of novel microRNAs with selective autophagy activity enabled multi-layered defence cascade against M. phaseolina in jute.     

动物成肌分化及非编码RNA调控研究进展

肌细胞分化密切关系到肉用动物的肌肉产量,也与人类的一系列疾病(如肌肉萎缩、心脏病等)密切相关。胚胎成肌分化期决定了肌纤维数量,是动物骨骼肌发育的关键时期。动物成肌分化及骨骼肌发育严格受各种细胞信号分子和转录因子调控,其中microRNA(miRNA)和lncRNA发挥着重要作用。本文从动物胚胎成肌分化及调控途径、卫星细胞的分化及调控、非编码RNA对肌肉形成的调控等方面进行综述,并展望了畜禽动物骨骼肌生长发育分子调控机制的研究方向,为提高畜禽肌肉产量与质量提供一定的分子理论参考。     

Relationship of microRNA-133a and TGF-β1 protein in myocardial tissues of spontaneously hypertensive rats

AIM：To observe the changes of microRNA-133a and transforming growth factor β1 (TGF-β1) protein in the myocardium of spontaneously hypertensive rats (SHR). METHODS：Male SHR (18 weeks old, n=12) and male Wistar-Kyoto rats (WKY, 18 weeks old, n=12) served as SHR group and control group, respectively. Caudal arterial blood pressure was detected by a noninvasive blood pressure measurement and analysis system. Myocardial collagen volume fraction (CVF) and perivascular collagen area ratio (PVCA) were determined by Masson staining. The level of miR-133a in the heart was detected by real-time quantitative PCR. The protein level of TGF-β1 in the heart was also analyzed by the methods of immunohistochemisty and Western blotting. RESULTS：Compared with control group, systolic and diastolic blood pressure, CVF and PVCA significantly increased, the expression of TGF-β1 protein was significantly up-regulated, and the level of miR-133a was significantly reduced in SHR group. In SHR group, the expression of miR-133a was decreased to (23.9±4.6)% in control group. A negative correlation between the levels of miR-133a and TGF-β1 protein in SHR group was observed (r=-0.791, P<0.01). CONCLUSION：The level of miR-133a is down-regulated along with the up-regulation of TGF-β1 protein expression and collagen synthesis in the myocardial tissues of SHR. miR-133a and TGF-β1 may be involved in myocardial fibrosis in SHR.     

Differential microRNA expression profile in laryngeal cancer and effect of miR-125a-5p on proliferation of laryngeal cancer cell line

AIM：To investigate the differential microRNA expression profiles between laryngeal cancer and adjacent normal laryngeal mucosa. METHODS：Forty two pairs of laryngeal cancer tissue and adjacent normal laryngeal mucosa tissue were collected. Ten pairs of samples were used for determining microRNA expression by the method of miRNA microarray chip. Data analysis was performed to find out the significant differential microRNA expression profile in laryngeal cancer, and the difference was verified by quantitative real-time PCR (qRT-PCR) analysis on another 32 pairs of samples. Methyl thiazolyl tetrazolium (MTT) assay and colony-forming assay were used to analyze the proliferation of Hep2 cells induced by miR-125a-5p. RESULTS：Both miRNA microarray and qRT-PCR showed that the expression of let-7f-5p, miR-10a-5p, miR-125a-5p, miR-144-3p, miR-195-5p and miR-203 was down-regulated in laryngeal cancer tissues. miR-125a-5p suppressed the proliferation of Hep2 cells. CONCLUSION：The results of microarray are accordant with those of qRT-PCR. Significant difference of miRNA expression profiles between laryngeal cancer and adjacent normal laryngeal mucosa indicates that miRNAs may play a role in carcinogenesis and progression of laryngeal cancer. miR-125a-5p inhibits the proliferation of Hep2 cell, indicating a novel therapeutic target against laryngeal cancer.     

Role of up-regulated microRNA145 in viability,apoptosis, invasion and metastasis of hepatoma cells

AIM: To investigate the effects of microRNA145 (miRNA145) on the viability, apoptosis, invasion and metastasis of hepatoma HepG2 cells. METHODS: HepG2 cells were randomly allocated into 3 groups: blank control group, empty mimic transfected group and miRNA145 mimic transfected group. Under the induction of Lipofectamine^TM 2000, the recombinant was transfected into HepG2 cells. After transfection, the expression level of miRNA145 was detected by real-time PCR. The protein level of N-cadherin and the mRNA expression levels of miRNA145 and N-cadherin were detected by Western blot and real-time PCR. The cell viability was detected by MTS assay. The cell cycle and apoptosis were analyzed by flow cytometry. Invasion and metastasis were detected by Transwell assay. RESULTS: Compared with negative control, miRNA145 expression was up-regulated significantly, while the expression of N-cadherin was down-regulated significantly. Meanwhile, the cell viability, cell cycle, apoptosis, invasion and metastasis of hepatoma HepG2 cells were all significantly inhibited (P<0.05). CONCLUSION: miRNA145 dramatically inhibits viability, apoptosis, invasion and metastasis of hepatoma cells.     

SNPs in APOBEC3 cytosine deaminases and their association with Visna/Maedi disease progression

The Apolipoprotein B mRNA-editing catalytic polypeptide-like 3 (APOBEC3) genes are able to inhibit the replication of a wide range of exogenous retroviruses, as well as endogenous retroviruses and retrotransposons.Three APOBEC3 genes, named APOBEC3Z1, APOBEC3Z2 and APOBEC3Z3, have been described in sheep.In this work the three genes have been screened in order to identify polymorphisms. No polymorphism was detected for the A3Z2 and A3Z3 genes but 16 SNPs and a 3-bp deletion were found in the A3Z1 gene. A thermoestability prediction analysis was applied to the detected amino acidic SNPs by three different programs. This analysis revealed a number of polymorphisms that could affect the protein stability. The SNPs of the 3′UTR were tested to detect alterations on the predicted microRNA target sites. Two new microRNA target sites were discovered for one of the alleles.Two SNPs were selected for association studies in relation with the retroviral disease Visna/Maedi in Latxa and Assaf sheep breeds. Although association analyses resulted unconclusive, probably due to the unsuitability of the SNP allele frequency distribution of the selected polymorphisms in the analyzed breeds, these genes remain good candidates for association studies.     

Differential expression of microRNA-206 in skeletal muscle of female Piedmontese and Friesian cattle

The double-muscle phenotype is an inherited condition in Piedmontese cattle traced to a point mutation in the myostatin gene. To investigate the potential role of muscle-specific miRNAs in determining muscle development in cattle, this study examined the patterns of expression of microRNAs (miRNA-1) and miRNA-206 in Piedmontese and Friesian cattle according to phenotype and sex. There were no significant differences in miRNA-1 expression between different muscle phenotypes, sexes or breeds, whereas there was significantly higher expression of miRNA-206 in female Piedmontese compared with female Friesian cattle.     

小分子microRNA及其生物学特征

MicroRNA(miRNA)是机体内源性表达的一类长20～24nt的非编码RNA调控因子,在动植物细胞中导致转录后基因沉默。近来发现miRNA家族成员众多,广泛地存在于各种真核细胞中,对诸如生长、发育、分化、死亡等生物过程进行调控。文章就其发现、形成及功能加以综述。     

初情期山羊胸腺microRNAs的筛选

为分析初情期前和初情期雌性山羊胸腺中MicroRNA的差异表达情况,并探讨其在山羊初情期启动过程中所起的作用.对初情期前(n = 3)和初情期(n = 3)山羊的胸腺组织进行了 Solexa测序,采用GO富集和KEGG分析评估差异miRNA靶基因.在初情期前和初情期的山羊胸腺组织中检测到538个miRNA,其中64个miRNA显著差异表达.与初情期前的山羊胸腺样本相比,初情期胸腺样本中存在19个上调基因和45个下调基因.KEGG通路富集分析显示,差异miRNA靶基因主要富集在苯丙氨酸,酪氨酸和色氨酸的生物合成,丁螺菌素和新霉素的生物合成等通路以及胆碱能突触和过氧化物酶体增殖物激活受体(PPAR)等与初情期启动相关信号途径.结果提示胸腺中miRNA参与山羊初情期启动.