Deep annotation of mouse iso-miR and iso-moR variation

Neuroscience Research Institute and Department of Cellular Molecular and Developmental Biology, University of California, Santa Barbara, CA 93106, USA.
Nucleic Acids Research (Impact Factor: 9.11). 03/2012; 40(13):5864-75. DOI: 10.1093/nar/gks247
Source: PubMed


With a dataset of more than 600 million small RNAs deeply sequenced from mouse hippocampal and staged sets of mouse cells that underwent reprogramming to induced pluripotent stem cells, we annotated the stem-loop precursors of the known miRNAs to identify isomoRs (miRNA-offset RNAs), loops, non-preferred strands, and guide strands. Products from both strands were readily detectable for most miRNAs. Changes in the dominant isomiR occurred among the cell types, as did switches of the preferred strand. The terminal nucleotide of the dominant isomiR aligned well with the dominant off-set sequence suggesting that Drosha cleavage generates most miRNA reads without terminal modification. Among the terminal modifications detected, most were non-templated mono- or di-nucleotide additions to the 3'-end. Based on the relative enrichment or depletion of specific nucleotide additions in an Ago-IP fraction there may be differential effects of these modifications on RISC loading. Sequence variation of the two strands at their cleavage sites suggested higher fidelity of Drosha than Dicer. These studies demonstrated multiple patterns of miRNA processing and considerable versatility in miRNA target selection.

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    • "3A; Additional file 1: Figure S4). This is consistent with the general observation that 3’-end sequence variation is more common than 5’-end variation [20, 32, 34, 39]. Within all mature miRNA reads, around 14% reads have undergone more than 1 nt trimming or degradation at the 3’-end. "
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    ABSTRACT: Background: microRNAs (miRNAs) are a highly abundant class of small noncoding regulatory RNAs that post-transcriptionally regulate gene expression in multicellular organisms. miRNAs are involved in a wide range of biological and physiological processes, including the regulation of host immune responses to microbial infections. Small-scale studies of miRNA expression in the malaria mosquito Anopheles gambiae have been reported, however no comprehensive analysis of miRNAs has been performed so far. Results: Using small RNA sequencing, we characterized de novo A. gambiae miRNA repertoire expressed in adult sugar- and blood-fed females. We provided transcriptional evidences for 123 miRNAs, including 58 newly identified miRNAs. Out of the newly described miRNAs, 19 miRNAs are homologs to known miRNAs in other insect species and 17 miRNAs share sequence similarity restricted to the seed sequence. The remaining 21 novel miRNAs displayed no obvious sequence homology with known miRNAs. Detailed bioinformatics analysis of the mature miRNAs revealed a sequence variation occurring at their 5'-end and leading to functional seed shifting in more than 5% of miRNAs. We also detected significant sequence heterogeneity at the 3'-ends of the mature miRNAs, mostly due to imprecise processing and post-transcriptional modifications. Comparative analysis of arm-switching events revealed the existence of species-specific production of dominant mature miRNAs induced by blood feeding in mosquitoes. We also identified new conserved and fragmented miRNA clusters and A. gambiae-specific miRNA gene duplication. Using miRNA expression profiling, we identified the differentially expressed miRNAs at an early time point after regular blood feeding and after infection with the rodent malaria parasite Plasmodium berghei. Significant changes were detected in the expression levels of 4 miRNAs in blood-fed mosquitoes, whereas 6 miRNAs were significantly upregulated after P. berghei infection. Conclusions: In the current study, we performed the first systematic analysis of miRNAs in A. gambiae. We provided new insights on mature miRNA sequence diversity and functional shifts in the mosquito miRNA evolution. We identified a set of the differentially expressed miRNAs that respond to normal and infectious blood meals. The extended set of Anopheles miRNAs and their isoforms provides a basis for further experimental studies of miRNA expression patterns and biological functions in A. gambiae.
    BMC Genomics 07/2014; 15(1):557. DOI:10.1186/1471-2164-15-557 · 3.99 Impact Factor
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    • "Processed reads were aligned to the mature Mus musculus miRNA database (, allowing one mismatch to occur between a read and the reference, due to isomiRs [19]. Sixty-six and 61.4% of the total reads from cochlear and vestibular sensory epithelia, respectively, were aligned to known mouse miRNAs, indicating a successful isolation of miRNAs. "
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    ABSTRACT: The mammalian inner ear contains sensory organs, the organ of Corti in the cochlea and cristae and maculae in the vestibule, with each comprised of patterned sensory epithelia that are responsible for hearing and balance. The development, cell fate, patterning, and innervation of both the sensory and nonsensory regions of the inner ear are governed by tight regulation involving, among others, transcription factors and microRNAs (miRNAs). In humans, mutations in specific miRNA genes are associated with hearing loss. In mice, experimental reduction or mutations of miRNAs in the inner ear leads to severe developmental and structural abnormalities. A comprehensive identification of miRNAs in the sensory epithelia and their gene targets will enable pathways of auditory and vestibular function to be defined. In this study, we used Next-Generation Sequencing (NGS) to identify the most prominent miRNAs in the inner ear and to define miRNA-target pairs that form pathways crucial for the function of the sensory epithelial cells. NGS of RNA from inner ear sensory epithelial cells led to the identification of 455 miRNAs in both cochlear and vestibular sensory epithelium, with 30 and 44 miRNAs found in only cochlea or vestibule, respectively. miR-6715-3p and miR-6715-5p were defined for the first time in the inner ear. Gene targets were identified for each of these miRNAs, including Arhgap12, a GTPase activating protein, for miR-6715-3p, implicating this miRNA in sensory hair cell bundle development, actin reorganization, cell adhesion and inner ear morphogenesis. This study provides a comprehensive atlas of miRNAs in the inner ear sensory epithelia. The results provide further support of the essential regulatory role of miRNAs in inner ear sensory epithelia and in regulating pathways that define development and growth of these cells.
    BMC Genomics 06/2014; 15(1):484. DOI:10.1186/1471-2164-15-484 · 3.99 Impact Factor
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    • "miRseq reads from these samples were mapped to well-authenticated miRNA precursors from miRBase.v16 using the small RNA pipeline from SOLiD, and only uniquely mapped reads were counted for miRNA profiling (Table S1; GEO accession number: GSE52608). Many hairpins had significant numbers of reads from both strands (Zhou et al., 2012). This data set contained a total of 766 miRNA precursors or 1,532 miRNA arms (5p and 3p), which were loaded to the edgeR package (Robinson et al., 2010). "
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    ABSTRACT: Major nonprimate-primate differences in corticogenesis include the dimensions, precursor lineages, and developmental timing of the germinal zones (GZs). microRNAs (miRNAs) of laser-dissected GZ compartments and cortical plate (CP) from embryonic E80 macaque visual cortex were deep sequenced. The CP and the GZ including ventricular zone (VZ) and outer and inner subcompartments of the outer subventricular zone (OSVZ) in area 17 displayed unique miRNA profiles. miRNAs present in primate, but absent in rodent, contributed disproportionately to the differential expression between GZ subregions. Prominent among the validated targets of these miRNAs were cell-cycle and neurogenesis regulators. Coevolution between the emergent miRNAs and their targets suggested that novel miRNAs became integrated into ancient gene circuitry to exert additional control over proliferation. We conclude that multiple cell-cycle regulatory events contribute to the emergence of primate-specific cortical features, including the OSVZ, generated enlarged supragranular layers, largely responsible for the increased primate cortex computational abilities.
    Neuron 02/2014; 81(6). DOI:10.1016/j.neuron.2014.01.017 · 15.05 Impact Factor
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