Regulatory Potential of Competing Endogenous RNAs in Myotonic Dystrophies.

Non-coding RNAs (ncRNAs) have been reported to be implicated in cell fate determination and various human diseases. All ncRNA molecules are emerging as key regulators of diverse cellular processes; however, little is known about the regulatory interaction among these various classes of RNAs. It has been proposed that the large-scale regulatory network across the whole transcriptome is mediated by competing endogenous RNA (ceRNA) activity attributed to both protein-coding and ncRNAs.

A potential role of extended simple sequence repeats in competing endogenous RNA crosstalk.

MicroRNA (miRNA)-mediated crosstalk between coding and non-coding RNAs of various types is known as the competing endogenous RNA (ceRNA) concept. Here, we propose that there is a specific variant of the ceRNA language that takes advantage of simple sequence repeat (SSR) wording. We applied bioinformatics tools to identify human transcripts that may be regarded as repeat-associated ceRNAs (raceRNAs).

Cooperation meets competition in microRNA-mediated DMPK transcript regulation.

The fundamental role of microRNAs (miRNAs) in the regulation of gene expression has been well-established, but many miRNA-driven regulatory mechanisms remain elusive. In the present study, we demonstrate that miRNAs regulate the expression of DMPK, the gene mutated in myotonic dystrophy type 1 (DM1), and we provide insight regarding the concerted effect of the miRNAs on the DMPK target. Specifically, we examined the binding of several miRNAs to the DMPK 3' UTR using luciferase assays.

In Vitro Expansion of CAG, CAA, and Mixed CAG/CAA Repeats.

Polyglutamine diseases, including Huntington's disease and a number of spinocerebellar ataxias, are caused by expanded CAG repeats that are located in translated sequences of individual, functionally-unrelated genes. Only mutant proteins containing polyglutamine expansions have long been thought to be pathogenic, but recent evidence has implicated mutant transcripts containing long CAG repeats in pathogenic processes. The presence of two pathogenic factors prompted us to attempt to distinguish the effects triggered by mutant protein from those caused by mutant RNA in cellular models of polyglutamine diseases.

Current understanding of the role of microRNAs in spinocerebellar ataxias.

The number of studies highlighting the role of microRNAs (miRNAs) in human physiology and diseases is growing, but many miRNA-driven regulatory mechanisms remain elusive. A proper understanding of the exact functions of individual miRNAs and their interaction with specific targets is vitally important because such knowledge might help cure diseases for which no effective treatment currently exists. Herein, we present current views on the role of the miRNA-mediated regulation of gene expression in the case of select spinocerebellar ataxias (SCAs) and their potential involvement in the pathogenesis of these diseases.

Regulation of huntingtin gene expression by miRNA-137, -214, -148a, and their respective isomiRs.

With the advent of deep sequencing technology, a variety of miRNA length and sequence variants, termed isomiRNAs (isomiRs), have been discovered. However, the functional roles of these commonly detected isomiRs remain unknown. In this paper, we demonstrated that miRNAs regulate the expression of the HTT gene, whose mutation leads to Huntington's disease (HD), a hereditary degenerative disorder.

The role of Dicer protein partners in the processing of microRNA precursors.

One of the cellular functions of the ribonuclease Dicer is to process microRNA precursors (pre-miRNAs) into mature microRNAs (miRNAs). Human Dicer performs this function in cooperation with its protein partners, AGO2, PACT and TRBP. The exact role of these accessory proteins in Dicer activity is still poorly understood.

The role of the precursor structure in the biogenesis of microRNA.

The human genome contains more than 1,000 microRNA (miRNA) genes, which are transcribed mainly by RNA polymerase II. The canonical pathway of miRNA biogenesis includes the nuclear processing of primary transcripts (pri-miRNAs) by the ribonuclease Drosha and further cytoplasmic processing of pre-miRNAs by the ribonuclease Dicer. This review discusses the issue of miRNA end heterogeneity generated primarily by Drosha and Dicer cleavage and focuses on the structural aspects of the Dicer step of miRNA biogenesis.

Northern blotting analysis of microRNAs, their precursors and RNA interference triggers.

Background: Numerous microRNAs (miRNAs) have heterogeneous ends resulting from imprecise cleavages by processing nucleases and from various non-templated nucleotide additions. The scale of miRNA end-heterogeneity is best shown by deep sequencing data revealing not only the major miRNA variants but also those that occur in only minute amounts and are unlikely to be of functional importance. All RNA interference (RNAi) technology reagents that are expressed and processed in cells are also exposed to the same machinery generating end-heterogeneity of the released short interfering RNAs (siRNAs) or miRNA mimetics.

High-resolution northern blot for a reliable analysis of microRNAs and their precursors.

This protocol describes how to perform northern blot analyses to detect microRNAs and their precursors with single-nucleotide resolution, which is crucial for analyzing individual length variants and for evaluating relative quantities of unique microRNAs in cells. Northern blot analysis consists of resolving RNAs by gel electrophoresis, followed by transferring and fixing to nylon membranes as well as detecting by hybridization with radioactive probes. Earlier efforts to improve this method focused mainly on altering the sensitivity of short RNA detection.

Structural basis of microRNA length variety.

The biogenesis of human microRNAs (miRNAs) includes two RNA cleavage steps in which the activities of the RNases Drosha and Dicer are involved. miRNAs of diverse lengths are generated from different genes, and miRNAs that are heterogeneous in length are produced from a single miRNA gene. We determined the solution structures of many miRNA precursors and analysed the structural basis of miRNA length diversity using a new measure: the weighted average length of diced RNA (WALDI).

Prediction and preliminary validation of oncogene regulation by miRNAs.

Background: MicroRNAs (miRNAs) are one of the most abundant groups of regulatory genes in multicellular organisms, playing important roles in many fundamental cellular processes. More than four hundred miRNAs have been identified in humans and the deregulation of miRNA expression has been also shown in many cancers. Despite the postulated involvement of miRNAs in tumourigenesis, there are only a few examples where an oncogene or a tumour suppressor has been identified as a miRNA target.

Analysis of RNA silencing in agroinfiltrated leaves of Nicotiana benthamiana and Nicotiana tabacum.

In this study we analyse several aspects of cytoplasmic RNA silencing by agroinfiltration of DNA constructs encoding single- and double-stranded RNAs derived from a GFP transgene and from the endogenous Virp1 gene. Both types of inductors resulted after 2-4 days in much higher concentration of siRNAs in the agroinfiltrated zone than normally seen during systemic silencing. More specifically, infiltration of two transgene hairpin constructs resulted in elevated levels of siRNAs.