BK virus encoded microRNAs are present in blood of renal transplant recipients with BK viral nephropathy.

BK viral infection is an important cause of renal transplant dysfunction and failure. Current strategies utilize surveillance for infection with DNA polymerase chain reaction assays and modulation of immunosuppression. Many viruses including polyomaviruses encode microRNAs (miRNAs). We have detected BK virus (BKV) encoded miRNAs in the blood of infected renal transplant recipients, and see a strong correlation between BKV encoded miRNA and BKV DNA in blood and a relationship between levels of bkv-miR-B1-5p and the presence of biopsy-proven BK viral nephropathy. Further research is needed to determine whether the detection of this and other virally encoded miRNAs may be useful in the diagnosis of active viral replication.

MicroRNA-143 expression in dorsal root ganglion neurons.

The unpleasant sensory and emotional experience of pain is initiated by excitation of primary afferent nociceptive neurons. Nerve damage or inflammation induces changes in nociceptive DRG neurons which contribute to both peripheral and central sensitization of pain-sensitive pathways. Recently, blockade of microRNA synthesis has been found to modulate the response of nociceptive neurons to inflammatory stimuli. However, little is known about the contributions of individual miRNAs to painful conditions. We compared miRNA expression in mouse sensory neurons and focussed on the localisation and control of miR-143. Using miRNA-arrays we compared the microRNA expression profile of intact lumbar DRG with one-day-old DRG cultures and found that nine miRNAs including miR-143 showed lower expression levels in cultures. Subsequent RT-qPCR confirmed array data and in-situ hybridisation localised miR-143 in the cytosol of sensory DRG neurons in situ and in vitro. Analysis of microbead-enriched neuron cultures showed significantly higher expression levels of miR-143 in isolectin B4 (I-B4) binding sensory neurons compared with neurons in the I-B4 negative flow-through fraction. In animal models of peripheral inflammation (injection of Complete Freund's Adjuvant, CFA) and nerve damage (transection of the sciatic nerve), we found that expression levels of miR-143 were significantly lower in DRGs ipsilateral to CFA injection or after nerve damage. Taken together, our data demonstrate for the first time miR-143 expression in nociceptive neurons. Since expression levels of miR-143 were higher in I-B4 positive neurons and declined in response to inflammation but not axotomy, miR-143 could selectively contribute to mRNA regulation in specific populations of nociceptors.

MicroRNA profiling of Barrett's oesophagus and oesophageal adenocarcinoma.

BACKGROUND: The genetic changes that drive metaplastic progression from squamous oesophageal mucosa toward intestinal metaplasia and adenocarcinoma are unclear. The aberrant expression of microRNAs (miRNAs) is involved in the development of cancer. This study examined whether miRNAs play a role in the development of oesophageal adenocarcinoma. METHODS: RNA was extracted from mucosa of normal oesophageal squamous epithelium, normal gastric epithelium, Barrett's oesophagus with intestinal metaplasia and oesophageal adenocarcinoma obtained from 16 individuals. Expression profiles of 377 human miRNAs were determined by microarray analysis and selected miRNAs were analysed further using real-time reverse transcription-polymerase chain reaction (RT-PCR) in tissues from 32 individuals. RESULTS: Microarray analyses identified 44 miRNAs likely to have altered expression between various mucosal samples. Of these, miR-21, miR-143, miR-145, miR-194, miR-203, miR-205 and miR-215 were chosen for validation by real-time RT-PCR. Tissue-specific expression profiles were observed, with miR-21, miR-143, miR-145, miR-194 and miR-215 significantly upregulated in columnar tissues compared with normal squamous epithelium. Expression of miR-143, miR-145 and miR-215 was lower in oesophageal adenocarcinoma than in Barrett's oesophagus. Levels of miR-203 and miR-205 were high in normal squamous epithelium and low in columnar epithelia. MiR-205 levels were lower in gastric epithelium than in both Barrett's oesophagus and adenocarcinoma. CONCLUSION: Expression of miRNA might define disease states in oesophageal epithelium. Dysregulation of specific miRNAs could contribute to metaplastic and neoplastic processes in the oesophageal mucosa.

MicroRNAs and cancer.

BACKGROUND: MicroRNAs (miRNAs) are small sequences of RNA, 21 to 22 nucleotides long, that have been discovered recently. They are produced from areas of the human genome that were previously thought to have no function. These sequences now appear to be important in the regulation of many fundamental processes. Evidence has recently emerged that deregulated miRNA activity is associated with human cancers. METHODS: The English literature was searched using PubMed for publications relevant to miRNAs and cancer. Relevant references from identified publications were also sourced. These publications were reviewed to identify existing evidence for the role of miRNAs in cancer. RESULTS: miRNAs inhibit the translation of mRNA from many target genes involved in cancer development. This leads to changes in the levels of protein encoded by these target genes and drives the development of cancer. The genes that produce miRNAs are frequently located in regions of the genome that are either lost, or amplified, in cancer cells. CONCLUSION: Determination of the miRNA expression profile in cancer tissues should lead to a better understanding of the genetic pathways involved in tumour development.

Quantitative evaluation of Escherichia coli host strains for tolerance to cytosine methylation in plasmid and phage recombinants.

Many strains of E. coli K12 restrict DNA containing cytosine methylation such as that present in plant and animal genomes. Such restriction can severely inhibit the efficiency of cloning genomic DNAs. We have quantitatively evaluated a total of 39 E. coli strains for their tolerance to cytosine methylation in phage and plasmid cloning systems. Quantitative estimations of relative tolerance to methylation for these strains are presented, together with the evaluation of the most promising strains in practical recombinant cloning situations. Host strains are recommended for different recombinant cloning requirements. These data also provide a rational basis for future construction of 'ideal' hosts combining optimal methylation tolerance with additional advantageous mutations.