Two less common human microRNAs miR-875 and miR-3144 target a conserved site of E6 oncogene in most high-risk human papillomavirus subtypes

Human papillomaviruses （HPVs） including high.risk （HR） and low-risk （LR） subtypes have distinguishable variation on both genotypes and phenotypes. The co- infection of multiple HR-HPVs, headed by HPV16, is common in cervical cancer in female. Recently accu- mulating reports have focused on the interaction be- tween virus and host, particularly the role of human microRNAs （miRNAs） in anti-viral defense by targeting viral genome. Here, we found a well-conserved target site of miRNAs in the genomes of most HR-HPVs, not LR-HPVs, by scanning all potential target sites of human miRNAs on 24 HPVs of unambiguous subtypes of risk. The site is targeted by two less common human miR- NAs, miR-875 and miR-3144, and is located in E6 onco- gene open reading frame （ORF） and overlap with the first alternative splice exon of viral early transcripts. In validation tests, miR-875 and miR-3144 were identified to suppress the target reporter activity markedly and inhibit the expression of both synthetically exogenous E6 and endogenous E6 oncogene. High level of two miRNAs can inhibit cell growth and promote apoptosisin HPV16-positive cervical cancer cells. This study pro- vides a promising common target of miRNAs for most HR-HPVs and highlights the effects of two low ex- pressed human miRNAs on tumour suppression.

MicroRNA-802和RAB23在前列腺癌中的表达水平及临床意义

目的探讨micro RNA-802(mi R-802)和RAB23在前列腺癌中的表达水平及临床意义。方法选取在海南省第三人民医院泌尿外科手术切除的前列腺癌标本60例,并选取其相对应癌旁组织作为对照,用免疫组织化学检测mi R-802和RAB23的表达水平,分析mi R-802和RAB23表达水平与前列腺癌患者临床病理参数间的关系及与前列腺癌患者预后。结果 miR-802和RAB23在前列腺癌组织中的阳性率分别为73.33%和70.00%,高于癌旁组织中的33.33%和28.33%(P <0.05)。mi R-802和RAB23表达水平与年龄和术前PSA无关(P>0.05);与Gleason评分相关,评分越高,表达率越低(P<0.05);与病理分期相关,表达高则病理分期早(P <0.05)。mi R-802和RAB23表达水平阴性组3年生存率及生存期均高于mi R-802和RAB23表达水平阳性组(P<0.05)。结论 miR-802和RAB23 mRNA在前列腺癌组织中的表达阳性率高于癌旁组织,阴性表达能获得较好的预后。

Identification of miR-802-5p and its involvement in type 2 diabetes mellitus

MicroRNAs(miRNA)are recently discovered endogenous,small noncoding RNAs(of 22 nucleotides)that play pivotal roles in gene regulation.They are involved in post-transcriptional control of gene expression.miRNAs are emerging as important regulators of cell proliferation,development,cancer formation,stress responses,cell death and physiological conditions.Increasing evidence has demonstrated the human miRNAs bind to their target mRNA sequences with perfect or near-perfect sequence complementarily.This provides a powerful strategy for discovering potential type 2 diabetes mellitus(T2DM)targets and gives the probability to exploit them for diagnostic and therapeutic causes.About 6%of the world population is affected by T2DM,and it is recognized as a global epidemic by the World Health Organization.At present there is no valid biomarker to control or manage T2DM.Therefore,the present study applied a mature sequence of miRNAs from publicly accessible databases to identify the miRNA from T2DM expressed sequence tags,and the results are detailed and discussed below.

Regulation of hepatic micro RNA expression by hepatocyte nuclear factor 4 alpha

AIMTo uncover the role of hepatocyte nuclear factor 4 alpha (HNF4α) in regulating hepatic expression of microRNAs. METHODSMicroarray and real-time PCR were used to determine hepatic expression of microRNAs in young-adult mice lacking Hnf4α expression in liver (Hnf4α-LivKO). Integrative genomics viewer software was used to analyze the public chromatin immunoprecipitation-sequencing datasets for DNA-binding of HNF4α, RNA polymerase-II, and histone modifications to loci of microRNAs in mouse liver and human hepatoma cells. Dual-luciferase reporter assay was conducted to determine effects of HNF4α on the promoters of mouse and human microRNAs as well as effects of microRNAs on the untranslated regions (3’UTR) of two genes in human hepatoma cells. RESULTSMicroarray data indicated that most microRNAs remained unaltered by Hnf4α deficiency in Hnf4α-LivKO mice. However, certain liver-predominant microRNAs were down-regulated similarly in young-adult male and female Hnf4α-LivKO mice. The down-regulation of miR-101, miR-192, miR-193a, miR-194, miR-215, miR-802, and miR-122 as well as induction of miR-34 and miR-29 in male Hnf4α-LivKO mice were confirmed by real-time PCR. Analysis of public chromatin immunoprecipitation-sequencing data indicates that HNF4α directly binds to the promoters of miR-101, miR-122, miR-194-2/miR-192 and miR-193, which is associated with histone marks of active transcription. Luciferase reporter assay showed that HNF4α markedly activated the promoters of mouse and human miR-101b/miR-101-2 and the miR-194/miR-192 cluster. Additionally, miR-192 and miR-194 significantly decreased activities of luciferase reporters for the 3’UTR of histone H3F3 and chromodomain helicase DNA binding protein 1 (CHD1), respectively, suggesting that miR-192 and miR-194 might be important in chromosome remodeling through directly targeting H3F3 and CHD1. CONCLUSIONHNF4α is essential for hepatic basal expression of a group of liver-enriched microRNAs, including miR-101, miR-192, miR-193a, miR-194 and miR-802, through which HNF4α may play a major role in the post-transcriptional regulation of gene expression and maintenance of the epigenome in liver.

Amphibian-derived peptide homodimer OA-GL17d promotes skin wound regeneration through the miR-663a/TGF-β1/Smad axis

Background:Amphibian-derived peptides exhibit considerable potential in the discovery and development of new therapeutic interventions for clinically challenging chronic skin wounds.MicroRNAs(miRNAs)are also considered promising targets for the development of effective therapies against skin wounds.However,further research in this field is anticipated.This study aims to identify and provide a new peptide drug candidate,as well as to explore the underlying miRNA mechanisms and possible miRNA drug target for skin wound healing.Methods:A combination of Edman degradation,mass spectrometry and cDNA cloning were adopted to determine the amino acid sequence of a peptide thatwas fractionated from the secretion of Odorrana andersonii frog skin using gel-filtration and reversed-phase high-performance liquid chromatography.The toxicity of the peptide was evaluated by Calcein-AM/propidium iodide(PI)double staining against human keratinocytes(HaCaT cells),hemolytic activity against mice blood cells and acute toxicity against mice.The stability of the peptide in plasma was also evaluated.The prohealing potency of the peptide was determined by MTS,scratch healing and a Transwell experiment against HaCaT cells,full-thickness injury wounds and scald wounds in the dorsal skin of mice.miRNA transcriptome sequencing analysis,enzyme-linked immunosorbent assay,real-time polymerase chain reaction and western blotting were performed to explore the molecular mechanisms.Results:A novel peptide homodimer(named OA-GL17d)that contains a disulfide bond between the 16th cysteine residue of the peptide monomer and the sequence‘GLFKWHPRCGEEQSMWT’was identified.Analysis showed that OA-GL17d exhibited no hemolytic activity or acute toxicity,but effectively promoted keratinocyte proliferation and migration and strongly stimulated the repair of full-thickness injury wounds and scald wounds in the dorsal skin of mice.Mechanistically,OA-GL17d decreased the level of miR-663a to increase the level of transforming growth factor-β1(TGF-β1)and activate the subsequent TGF-β1/Smad signaling pathway,thereby resulting in accelerated skin wound re-epithelialization and granular tissue formation.Conclusions:Our results suggest that OA-GL17d is a new peptide drug candidate for skin wound repair.This study emphasizes the importance of exogenous peptides as molecular probes for exploring competing endogenous RNA mechanisms and indicates that miR-663a may be an effective target for promoting skin repair.