An atlas of DNA methylomes in porcine adipose and muscle tissues

It is evident that epigenetic factors, especially DNA methylation, have essential roles in obesity development. Here, using pig as a model, we investigate the systematic association between DNA methylation and obesity. We sample eight variant adipose and two distinct skeletal muscle tissues from three pig breeds living within comparable environments but displaying distinct fat level. We generate 1,381 Gb of sequence data from 180 methylated DNA immunoprecipitation libraries, and provide a genome-wide DNA methylation map as well as a gene expression map for adipose and muscle studies. The analysis shows global similarity and difference among breeds, sexes and anatomic locations, and identifies the differentially methylated regions. The differentially methylated regions in promoters are highly associated with obesity development via expression repression of both known obesity-related genes and novel genes. This comprehensive map provides a solid basis for exploring epigenetic mechanisms of adipose deposition and muscle growth

MicroRNAome of Porcine Pre- and Postnatal Development

The domestic pig is of enormous agricultural significance and valuable models for many human diseases. Information concerning the pig microRNAome (miRNAome) has been long overdue and elucidation of this information will permit an atlas of microRNA (miRNA) regulation functions and networks to be constructed. Here we performed a comprehensive search for porcine miRNAs on ten small RNA sequencing libraries prepared from a mixture of tissues obtained during the entire pig lifetime, from the fetal period through adulthood. The sequencing results were analyzed using mammalian miRNAs, the precursor hairpins (pre-miRNAs) and the first release of the high-coverage porcine genome assembly (Sscrofa9, April 2009) and the available expressed sequence tag (EST) sequences. Our results extend the repertoire of pig miRNAome to 867 pre-miRNAs (623 with genomic coordinates) encoding for 1,004 miRNAs, of which 777 are unique. We preformed real-time quantitative PCR (q-PCR) experiments for selected 30 miRNAs in 47 tissue-specific samples and found agreement between the sequencing and q-PCR data. This broad survey provides detailed information about multiple variants of mature sequences, precursors, chromosomal organization, development-specific expression, and conservation patterns. Our data mining produced a broad view of the pig miRNAome, consisting of miRNAs and isomiRs and a wealth of information of pig miRNA characteristics. These results are prelude to the advancement in pig biology as well the use of pigs as model organism for human biological and biomedical studies

Microarray and degradome sequencing reveal microRNA differential expression profiles and their targets in Pinellia pedatisecta.

MicroRNAs (miRNAs) are endogenous small non-coding RNAs which play a critical role in gene regulation in plants. Pinelliapedatisecta is one of the most important herbs in traditional Chinese medicine, but there are no microRNAs of Pinelliapedatisecta were deposited in miRBase and the research of the related miRNA biological functions is still insufficient. To detect Pinelliapedatisecta miRNAs and discover their expression difference with Pinelliaternata, we carried out a microarray profiling. A total of 101 miRNAs belonging to 22 miRNA families were detected both in Pinelliapedatisecta and Pinelliaternata respectively, among them 21 miRNAs showed their differentially expression. GO (gene ontology) term enrichment analysis of the target genes of differential expression miRNAs reveal that these miRNAs mainly affect the reproduction, transcription factor activity and plant developmental process. To elucidate the target function of miRNAs, we constructed a degradome library from Pinellia pedatisecta leaf. The result showed that a total of 18 transcript were identified as targets of miRNAs and further analysis indicated that miR156 and miR529 may function together to repress SPL14

The homology of identified miRNAs in other plant species.

<p>Values on Y axis indicate the number of homological miRNA between <i>Pinellia</i> and other plant species.</p

Validation and expression analysis of miRNAs by qRT-PCR (<i>P</i>.<i>pedatisecta</i>/<i>P</i>. <i>ternate</i>).

<p>Blue indicates the miRNA relative expression generated from microarray; Red indicates the miRNA relative expression tested by qRT-PCR. The 5.8s rRNA was chosen as the endogenous control. The error bars indicate the standard deviations obtained from three independent experiments of qRT-PCR.</p

Differential-expression microRNA-mediated regulatory networks.

<p>(A) Network mediated by high expression microRNAs in <i>P</i><i>. pedatisecta</i>. (B) Network mediated by low expression microRNAs in <i>P</i><i>. pedatisecta</i>. The predicted regulatory relationships between miRNAs and targets were denoted by blank lines. The targets of blue nodes were annotated at the bottom and all the networks were constructed using Cytoscape [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0075978#B35" target="_blank">35</a>].</p

Cleavage analyses with respect to SPL14 targeted by miR529 and miR156.

<p>(1) SPL14 mRNA(LOC_Os08g39890) aligned with miR529 and miR156. (2) miR529 and miR156 sequence alignments. *means homology sequence(3). Cleavage sites of SPL14 transcript by miR156 were determined by Jiao et al using 5’ RACE assay. The number of clones which represent the cleavage frequencies is indicated above the sequence.</p

Identified miRNA targets by degradome sequencing are presented in the form of t-plots.

<p>The T-plots which were referred to as ‘target plots’ by German et al indicated the distribution of the degradome tags along the sequences of targets (top). The alignment of mRNA: miRNA show the matching degree between miRNA and its target sequence (bottom). The two dots indicate matched RNA base pairs; one dot indicates a GU mismatch. The red spot and arrow represents the sliced target site. The y-axis measures the intensity of the cleavage signal, and the x-axis indicates the position of the cleavage signal on a specific transcript. (A) Regulation of SBP-box gene family by miR156 and miR157 family identified by degradome sequencing. (B) Different target genes cleaved by the miR529 family.</p

Number of miRNA members in each family which commonly identified from <i>P</i><i>. pedatisecta</i> and <i>P</i><i>. ternate</i> by microarray.

<p>Number of miRNA members in each family which commonly identified from <i>P</i><i>. pedatisecta</i> and <i>P</i><i>. ternate</i> by microarray.</p