Finding motifs in miRNA sequences

Recently, the emergence of non-coding miRNA has attracted biology and computer researchers. miRNA plays an important role in regulation of genes. Finding motifs in RNA is one of important topics. In our work, we attempt to find motifs in mature miRNA from combinations ranging from two to ten nucleotides. Interestingly, we have found several motifs only appear in mature miRNA but not appear in other regions of primary miRNA sequences taken from latest miRNA datasets. The findings of our investigation may help in the building model to predict all possible miRNAs in genomes

Visualising single molecules of HIV-1 and miRNA nucleic acids

Background
The scarcity of certain nucleic acid species and the small size of target sequences such as miRNA, impose a significant barrier to subcellular visualization and present a major challenge to cell biologists. Here, we offer a generic and highly sensitive visualization approach (oligo fluorescent in situ hybridization, O-FISH) that can be used to detect such nucleic acids using a single-oligonucleotide probe of 19–26 nucleotides in length.

Results
We used O-FISH to visualize miR146a in human and avian cells. Furthermore, we reveal the sensitivity of O-FISH detection by using a HIV-1 model system to show that as little as 1–2 copies of nucleic acids can be detected in a single cell. We were able to discern newly synthesized viral cDNA and, moreover, observed that certain HIV RNA sequences are only transiently available for O-FISH detection.

Conclusions
Taken together, these results suggest that the O-FISH method can potentially be used for in situ probing of, as few as, 1–2 copies of nucleic acid and, additionally, to visualize small RNA such as miRNA. We further propose that the O-FISH method could be extended to understand viral function by probing newly transcribed viral intermediates; and discern the localisation of nucleic acids of interest. Additionally, interrogating the conformation and structure of a particular nucleic acid in situ might also be possible, based on the accessibility of a target sequence.

Differential temporal expression of milk miRNA during the lactation cycle of the marsupial tammar wallaby (Macropus eugenii)

Lactation is a key aspect of mammalian evolution for adaptation of various reproductive strategies along different mammalian lineages. Marsupials, such as tammar wallaby, adopted a short gestation and a relatively long lactation cycle, the newborn is immature at birth and significant development occurs postnatally during lactation. Continuous changes of tammar milk composition may contribute to development and immune protection of pouch young. Here, in order to address the putative contribution of newly identified secretory milk miRNA in these processes, high throughput sequencing of miRNAs collected from tammar milk at different time points of lactation was conducted. A comparative analysis was performed to find distribution of miRNA in milk and blood serum of lactating wallaby.

miRNA modulation of SOCS1 using an influenza A virus delivery system

Difficulties associated with efficient delivery and targeting of miRNAs to cells is hampering the real world application of miRNA technology. This study utilized an influenza A-based delivery system to express miR-155 in order to knockdown SOCS1 mRNA. Using qPCR and dual luciferase technology we show that miR-155 delivery resulted in a significant increase in cellular miR-155 which facilitated a downregulation of SOCS1 gene expression and a functional increase in IL-6 and IFN-β cytokines.

Blocking the maturation of OncomiRNAs using pri-miRNA-17∼92 aptamer in retinoblastoma

The miR-17∼92. or oncomiR-1, cluster encodes oncogenic microRNAs (miRNAs), and it also promotes retinoblastoma (RB) tumor formation. Antagomir and miRNA mimics based approaches are widely tried against oncogenic and tumor suppressive miRNAs. Other methods for targeting cancer related miRNAs are still under development. In the current study, we focused on the pri-miRNA-17∼92 aptamer (pri-apt), which can potentially replace the mix of five antagomirs by one aptamer that function to abrogate the maturation of miR-17, miR-18a, and miR-19b (P<0.05) for targeting RB. We used RB cell lines WERI-Rb1 and Y79 as an in vitro model. Cellular changes upon transfecting the pri-apt led to S-phase arrest in WERI-Rb1 cells and onset of apoptosis in both Y79 and WERI-Rb1 cell lines. There was increased cytotoxicity as measured by lactate dehydrogenase activity in pri-apt treated Y79 cells (P<0.05), and significant inhibition of cell proliferation was observed in both of the cell lines. Thus we showed the antiproliferative property of pri-apt in RB cell lines, which can be readily modified by developing appropriate vectors for the delivery of the aptamer specifically to cancer cells.

Finding edging genes from microarray data

Motivation: A set of genes and their gene expression levels are used to classify disease and normal tissues. Due to the massive number of genes in microarray, there are a large number of edges to divide different classes of genes in microarray space. The edging genes (EGs) can be co-regulated genes, they can also be on the same pathway or deregulated by the same non-coding genes, such as siRNA or miRNA. Every gene in EGs is vital for identifying a tissue's class. The changing in one EG's gene expression may cause a tissue alteration from normal to disease and vice versa. Finding EGs is of biological importance. In this work, we propose an algorithm to effectively find these EGs.

Result: We tested our algorithm with five microarray datasets. The results are compared with the border-based algorithm which was used to find gene groups and subsequently divide different classes of tissues. Our algorithm finds a significantly larger amount of EGs than does the border-based algorithm. As our algorithm prunes irrelevant patterns at earlier stages, time and space complexities are much less prevalent than in the border-based algorithm.

Changes in the composition of the extracellular matrix in patellar tendinopathy

Objective: To compare the chemical levels and mRNA expression of proteoglycan and collagen in normal human patellar tendons and tendons exhibiting chronic overuse tendinopathy.

Methods: Sulfated glycosaminoglycan and hydroxyproline content were investigated by spectrophotometric measurement using papain-digested samples. Deglycosylated proteoglycan core proteins were analysed by Western blot using specific antibodies. Total mRNA isolated from samples of frozen tendons was assayed by relative quantitative RT-PCR for decorin, biglycan, fibromodulin, versican, aggrecan, and collagens Type I, II and III and normalised to glyceraldehyde-3-phosphate dehydrogenase.

Results: There was a significant increase in sulfated glycosaminoglycan content in pathologic tendons compared to normal. This was attributed to an increased deposition of the large aggregating proteoglycans versican and aggrecan and the small proteoglycans biglycan and fibromodulin, but not decorin. Aggrecan and versican were extensively degraded in both normal and pathologic tendons, biglycan was more fragmented in the pathologic tendons while predominantly intact fibromodulin and decorin were present in normal and pathologic tendons. There was a greater range in total collagen content but no change in the level of total collagen in pathologic tendons. There were no significant differences between the pathologic and normal tendon for all genes, however p values close to 0.05 indicated a trend in downregulation of Type I collagen and fibromodulin, and upregulation in versican and Type III genes in pathologic tissue.

Conclusion: The changes in proteoglycan and collagen levels observed in patellar tendinopathy appear to be primarily due to changes in the metabolic turnover of these macromolecules. Changes in the expression of these macromolecules may not play a major role in this process.

Viral RNA silencing suppressors (RSS) : novel strategy of viruses to ablate the host RNA interference (RNAi) defense system

Pathogenic viruses have developed a molecular defense arsenal for their survival by counteracting the host anti-viral system known as RNA interference (RNAi). Cellular RNAi, in addition to regulating gene expression through microRNAs, also serves as a barrier against invasive foreign nucleic acids. RNAi is conserved across the biological species, including plants, animals and invertebrates. Viruses in turn, have evolved mechanisms that can counteract this anti-viral defense of the host. Recent studies of mammalian viruses exhibiting RNA silencing suppressor (RSS) activity have further advanced our understanding of RNAi in terms of host–virus interactions. Viral proteins and non-coding viral RNAs can inhibit the RNAi (miRNA/siRNA) pathway through different mechanisms. Mammalian viruses having dsRNA-binding regions and GW/WG motifs appear to have a high chance of conferring RSS activity. Although, RSSs of plant and invertebrate viruses have been well characterized, mammalian viral RSSs still need in-depth investigations to present the concrete evidences supporting their RNAi ablation characteristics. The information presented in this review together with any perspective research should help to predict and identify the RSS activity-endowed new viral proteins that could be the potential targets for designing novel anti-viral therapeutics.

MicroRNA in human cancer and chronic inflammatory diseases

MicroRNAs (miRNAs) are the non-coding RNAs that act as post-translational regulators to their complimentary messenger RNAs (mRNA). Due to their specific gene silencing property, miRNAs have been implicated in a number of cellular and developmental processes. Also, it has been proposed that a particular set of miRNA spectrum is expressed only in a particular type of tissue. Many interesting findings related to the differential expression of miRNAs in various human diseases including several types of cancers, neurodegenerative diseases and metabolic diseases have been reported. Deregulation of miRNA expression in different types of human diseases and the roles various miRNAs play as tumour suppressors as well as oncogenes, suggest their contribution to cancer and/or in other disease development. These findings have possible implications in the development of diagnostics and/or therapeutics in human malignancies. In this review, we discuss various miRNAs that are differentially expressed in human chronic inflammatory diseases, neurodegenerative diseases, cancer and the further prospective development of miRNA based diagnostics and therapeutics.

Identifying co-regulating microrna groups

Background: Current miRNA target prediction tools have the common problem that their false positive rate is high. This renders identification of co-regulating groups of miRNAs and target genes unreliable. In this study, we describe a procedure to identify highly probable co-regulating miRNAs and the corresponding co-regulated gene groups. Our procedure involves a sequence of statistical tests: (1) identify genes that are highly probable miRNA targets; (2) determine for each such gene, the minimum number of miRNAs that co-regulate it with high probability; (3) find, for each such gene, the combination of the determined minimum size of miRNAs that co-regulate it with the lowest p-value; and (4) discover for each such combination of miRNAs, the group of genes that are co-regulated by these miRNAs with the lowest p-value computed based on GO term annotations of the genes.
Results: Our method identifies 4, 3 and 2-term miRNA groups that co-regulate gene groups of size at least 3 in human. Our result suggests some interesting hypothesis on the functional role of several miRNAs through a "guilt by association" reasoning. For example, miR-130, miR-19 and miR-101 are known neurodegenerative diseases associated miRNAs. Our 3-term miRNA table shows that miR-130/19/101 form a co-regulating group of rank 22 (p-value =1.16 × 10^-2). Since miR-144 is co-regulating with miR-130, miR-19 and miR-101 of rank 4 (p-value = 1.16 × 10^-2) in our 4-term miRNA table, this suggests hsa-miR-144 may be neurodegenerative diseases related miRNA. Conclusions: This work identifies highly probable co-regulating miRNAs, which are refined from the prediction by computational tools using (1) signal-to-noise ratio to get high accurate regulating miRNAs for every gene, and (2) Gene Ontology to obtain functional related co-regulating miRNA groups. Our result has partly been supported by biological experiments. Based on prediction by TargetScanS, we found highly probable target gene groups in the Supplementary Information. This result might help biologists to find small set of miRNAs for genes of interest rather than huge amount of miRNA set.