The species Brassica rapa includes various vegetable crops. Production of these vegetable crops is usually impaired by heat stress. Some microRNAs (miRNAs) in Arabidopsis have been considered to mediate gene silencing in plant response to abiotic stress. However, it remains unknown whether or what miRNAs play a role in heat resistance of B. rapa. To identify genomewide conserved and novel miRNAs that are responsive to heat stress in B. rapa, we defined temperature thresholds of non-heading Chinese cabbage (B. rapa ssp. chinensis) and constructed small RNA libraries from the seedlings that had been exposed to high temperature (46 °C) for 1 h. By deep sequencing and data analysis, we selected a series of conserved and novel miRNAs that responded to heat stress. In total, Chinese cabbage shares at least 35 conserved miRNA families with Arabidopsis thaliana. Among them, five miRNA families were responsive to heat stress. Northern hybridization and real-time PCR showed that the conserved miRNAs bra-miR398a and bra-miR398b were heat-inhibitive and guided heat response of their target gene, BracCSD1; and bra-miR156h and bra-miR156g were heat-induced and its putative target BracSPL2 was down-regulated. According to the criteria of miRNA and miRNA* that form a duplex, 21 novel miRNAs belonging to 19 miRNA families were predicted. Of these, four were identified to be heat-responsive by Northern blotting and/or expression analysis of the putative targets. The two novel miRNAs bra-miR1885b.3 and bra-miR5718 negatively regulated their putative target genes. 5′-Rapid amplification of cDNA ends PCR indicated that three novel miRNAs cleaved the transcripts of their target genes where their precursors may have evolved from. These results broaden our perspective on the important role of miRNA in plant responses to heat.

BackgroundNon-coding small RNAs play critical roles in various cellular processes in a wide spectrum of eukaryotic organisms. Their responses to abiotic stress have become a popular topic of economic and scientific importance in biological research. Several studies in recent years have reported a small number of non-coding small RNAs that map to chloroplast genomes. However, it remains uncertain whether small RNAs are generated from chloroplast genome and how they respond to environmental stress, such as high temperature. Chinese cabbage is an important vegetable crop, and heat stress usually causes great losses in yields and quality. Under heat stress, the leaves become etiolated due to the disruption and disassembly of chloroplasts. In an attempt to determine the heat-responsive small RNAs in chloroplast genome of Chinese cabbage, we carried out deep sequencing, using heat-treated samples, and analysed the proportion of small RNAs that were matched to chloroplast genome.ResultsDeep sequencing provided evidence that a novel subset of small RNAs were derived from the chloroplast genome of Chinese cabbage. The chloroplast small RNAs (csRNAs) include those derived from mRNA, rRNA, tRNA and intergenic RNA. The rRNA-derived csRNAs were preferentially located at the 3'-ends of the rRNAs, while the tRNA-derived csRNAs were mainly located at 5'-termini of the tRNAs. After heat treatment, the abundance of csRNAs decreased in seedlings, except those of 24 nt in length. The novel heat-responsive csRNAs and their locations in the chloroplast were verified by Northern blotting. The regulation of some csRNAs to the putative target genes were identified by real-time PCR. Our results reveal that high temperature suppresses the production of some csRNAs, which have potential roles in transcriptional or post-transcriptional regulation.ConclusionsIn addition to nucleus, the chloroplast is another important organelle that generates a number of small RNAs. Many members of csRNA families are highly sensitive to heat stress. Some csRNAs respond to heat stress by silencing target genes. We suggest that proper temperature is important for production of chloroplast small RNAs, which are associated with plant resistance to abiotic stress.

Background The majority of established prostate cancer risk-associated Single Nucleotide Polymorphisms (SNPs) identified from genome-wide association studies do not fall into protein coding regions. Therefore, the mechanisms by which these SNPs affect prostate cancer risk remain unclear. Here, we used a series of bioinformatic tools and databases to provide possible molecular insights into the actions of risk SNPs. Methodology/Principal Findings We performed a comprehensive assessment of the potential functional impact of 33 SNPs that were identified and confirmed as associated with PCa risk in previous studies. For these 33 SNPs and additional SNPs in Linkage Disequilibrium (LD) (r2 ≥ 0.5), we first mapped them to genomic functional annotation databases, including the Encyclopedia of DNA Elements (ENCODE), eleven genomic regulatory elements databases defined by the University of California Santa Cruz (UCSC) table browser, and Androgen Receptor (AR) binding sites defined by a ChIP-chip technique. Enrichment analysis was then carried out to assess whether the risk SNP blocks were enriched in the various annotation sets. Risk SNP blocks were significantly enriched over that expected by chance in two annotation sets, including AR binding sites (p=0.003), and FoxA1 binding sites (p=0.05). About one third of the 33 risk SNP blocks are located within AR binding regions. Conclusions/Significance The significant enrichment of risk SNPs in AR binding sites may suggest a potential molecular mechanism for these SNPs in prostate cancer initiation, and provide guidance for future functional studies.

Background Genome-wide association studies (GWAS) have identified approximately three dozen single nucleotide polymorphisms (SNPs) consistently associated with prostate cancer (PCa) risk. Despite the reproducibility of these associations, the molecular mechanism for most of these SNPs has not been well elaborated as most lie within non-coding regions of the genome. Androgens play a key role in prostate carcinogenesis. Recently, using ChIP-on-chip technology, 22,447 androgen receptor (AR) binding sites have been mapped throughout the genome, greatly expanding the genomic regions potentially involved in androgen-mediated activity. Methodology/Principal findings To test the hypothesis that sequence variants in AR binding sites are associated with PCa risk, we performed a systematic evaluation among two existing PCa GWAS cohorts; the Johns Hopkins Hospital and the Cancer Genetic Markers of Susceptibility (CGEMS) study population. We demonstrate that regions containing AR binding sites are significantly enriched for PCa risk-associated SNPs, i.e. more than expected by chance alone. In addition, compared with the entire genome, these newly observed risk-associated SNPs in these regions are significantly more likely to overlap with established PCa risk-associated SNPs from previous GWAS. These results are consistent with our previous finding from a bioinformatics analysis that one-third of the 33 known PCa risk-associated SNPs discovered by GWAS are located in regions of the genome containing AR binding sites. Conclusions/Significance The results to date provide novel statistical evidence suggesting an androgen-mediated mechanism by which some PCa associated SNPs act to influence PCa risk. However, these results are hypothesis generating and ultimately warrant testing through in-depth molecular analyses.