Background: Cellular miRNAs play an important role in the regulation of gene expression in eukaryotes. Recently, miRNAs have also been shown to be able to target and inhibit viral gene expression. Computational predictions revealed earlier that the HIV-1 genome includes regions that may be potentially targeted by human miRNAs. Here we report the functionality of predicted miR-29a target site in the HIV-1 nef gene.

Actin is a major cytoskeletal protein in eukaryotes. Recent studies suggest more diverse functional roles for this protein. Actin mRNA is known to be localized to neuronal synapses and undergoes rapid deadenylation during early developmental stages. However, its 3′-untranslated region (UTR) is not characterized and there are no experimentally determined polyadenylation (polyA) sites in actin mRNA. We have found that the cytoplasmic β-actin (Actb) gene generates two alternative transcripts terminated at tandem polyA sites. We used 3′-RACE, EST end analysis and in situ hybridization to unambiguously establish the existence of two 3′-UTRs of varying length in Actb transcript in mouse neuronal cells. Further analyses showed that these two tandem polyA sites are used in a tissue-specific manner. Although the longer 3′-UTR was expressed at a relatively lower level, it conferred higher translational efficiency to the transcript. The longer transcript harbours a conserved mmu-miR-34a/34b-5p target site. Sequence-specific anti-miRNA molecule, mutations of the miRNA target region in the 3′-UTR resulted in reduced expression. The expression was restored by a mutant miRNA complementary to the mutated target region implying that miR-34 binding to Actb 3′-UTR up-regulates target gene expression. Heterogeneity of the Actb 3′-UTR could shed light on the mechanism of miRNA-mediated regulation of messages in neuronal cells.

Critical features of stem cells include anchoring within a niche and activation upon injury. Notch signaling maintains skeletal muscle satellite (stem) cell quiescence by inhibiting differentiation and inducing expression of extracellular components of the niche. However, the complete spectrum of how Notch safeguards quiescence is not well understood. Here, we perform Notch ChIPsequencing and small RNA sequencing in satellite cells and identify the Notch-induced microRNA-708, which is a mirtron that is highly expressed in quiescent cells and sharply downregulated in activated cells. We employ in vivo and ex vivo functional studies, in addition to live imaging, to show that miR-708 regulates quiescence and selfrenewal by antagonizing cell migration through targeting the transcripts of the focal-adhesionassociated protein Tensin3. Therefore, this study identifies a Notch-miR708-Tensin3 axis and suggests that Notch signaling can regulate satellite cell quiescence and transition to the activation state through dynamic regulation of the migratory machinery.(I) miR-708 knockdown using AntimiR-708 transfection of single EDL myofibers from Tg:Pax7-nGFP mice cultured for 72 hr and immunostained for GFP and Myogenin. Quantification of PAX7 + /MYOG -(21% versus 5%), PAX7 -/MYOG -(36% versus 15%), and PAX7 -/MYOG + (43% versus 79%) for Scramble and AntimiR-708 populations respectively; 4 mice/condition, n = 25 fibers, is shown. (J) Satellite cells isolated by FACS from control and Odz4-null mice, treated with Mimic or Control for 72 hr and stained for Myogenin. Quantification of Myogenin + cells; n = 3 mice/condition, R250 cells, is shown. Error bars, mean ± SD; two-sided unpaired t test; # p value: two-sided paired t test. Scale bars: 50 mm. See also Figure S1.

MicroRNAs (miRNAs) are small, endogenous, regulatory RNA molecules that can bind to partially complementary regions on target messenger RNAs and impede their expression or translation. We rationalized that miRNAs, being localized to the cytoplasm, will be maternally inherited during fertilization and may play a role in early development. Although Dicer is known to be essential for the transition from single-celled zygote to two-cell embryo, a direct role for miRNAs has not yet been demonstrated. We identified miRNAs with targets in zygotically expressed transcripts in Drosophila using a combination of transcriptome analysis and miRNA target prediction. We experimentally established that Drosophila miRNA dme-miR-34, the fly homologue of the cancer-related mammalian miRNA miR-34, involved in somatic-cell reprogramming and having critical role in early neuronal differentiation, is present in Drosophila embryos before initiation of zygotic transcription. We also show that the Drosophila miR-34 is dependent on maternal Dicer-1 for its expression in oocytes. Further, we show that miR-34 is also abundant in unfertilized oocytes of zebrafish. Its temporal expression profile during early development showed abundant expression in unfertilized oocytes that gradually decreased by 5 days post-fertilization (dpf). We find that knocking down the maternal, but not the zygotic, miR-34 led to developmental defects in the neuronal system during early embryonic development in zebrafish. Here, we report for the first time, the maternal inheritance of an miRNA involved in development of the neuronal system in a vertebrate model system.