A Global Identification and Analysis of Small Nucleolar RNAs and Possible Intermediate-Sized Non-Coding RNAs in Oryza sativa

Accumulating evidence suggests that non-coding RNAs （ncRNAs） are both widespread and functionally important in many eukaryotic organisms. In this study, we employed a special size fractionation and cDNA library construction method followed by 454 deep sequencing to systematically profile rice intermediate-size ncRNAs. Our analysis resulted in the identification of 1349 ncRNAs in total, including 754 novel ncRNAs of an unknown functional category. Chromosome distribution of all identified ncRNAs showed no strand bias, and displayed a pattern similar to that observed in protein-coding genes with few chromosome dependencies. More than half of the ncRNAs were centered around the plus-strand of the 5＇ and 3＇ termini of the coding regions. The majority of the novel ncRNAs were rice specific, while 78% of the small nucleolar RNAs （snoRNAs） were conserved. Tandem duplication drove the expansion of over half of the snoRNA gene families. Furthermore, 90% of the snoRNA candidates were shown to produce small RNAs between 20-30 nt, 80% of which were associated with ARGONAUT proteins generally, and AGOlb in particular. Overall, our findings provide a comprehensive view of an intermediate-size non-coding transcriptome in a monocot species, which will serve as a useful platform for an in-depth analysis of ncRNA functions.

Genomic Features and Regulatory Roles of Intermediate-Sized Non-Coding RNAs in Arabidopsis

ABSTRACT Recent advances in genome-wide techniques allowed the identification of thousands of non-coding RNAs with various sizes in eukaryotes, some of which have further been shown to serve important functions in many biologi- cal processes. However, in model plant Arabidopsis, novel intermediate-sized ncRNAs （im-ncRNAs） （50-300 nt） have very limited information. By using a modified isolation strategy combined with deep-sequencing technology, we identified 838 im-ncRNAs in Arabidopsis globally. More than half （58%） are new ncRNA species, mostly evolutionary divergent. Interestingly, annotated protein-coding genes with 5＇-UTR-derived novel im-ncRNAs tend to be highly expressed. For intergenic im-ncRNAs, their average abundances were comparable to mRNAs in seedlings, but subsets exhibited signifi- cantly lower expression in senescing leaves. Further, intergenic im-ncRNAs were regulated by similar genetic and epige- netic mechanisms to those of protein-coding genes, and some showed developmentally regulated expression patterns. Large-scale reverse genetic screening showed that the down-regulation of a number of im-ncRNAs resulted in either obvious molecular changes or abnormal developmental phenotypes in vivo, indicating the functional importance of im-ncRNAs in plant growth and development. Together, our results demonstrate that novel Arabidopsis im-ncRNAs are developmentally regulated and functional components discovered in the transcriptome.

Natural variation of H3K27me3 modification in two Arabidopsis accessions and their hybrid

Histone modifications affect gene expression, but the mechanism and biological consequence of natural variation in histone modifications remain unclear. Here, we generated genome-wide integrated maps of H3K27me3 modification and transcriptome for Col, C24 and their F1 hybrid. A total of 1,828 genomic regions showing variation in H3K27me3 modification between Col and C24 were identified, most of which were associated with genic regions. Natural variation of H3K27me3 modification between parents could result in aUelic bias of H3K27me3 in hybrids. Furthermore, we found that H3K27me3 variation between Col and C24 was negatively correlated with gene expression differences between two accessions, espe- cially with those arising from the cis-effect. Importantly, mutation of CLF, an Arabidopsis methyltransferase for H3K27,altered gene expression patterns between the parents. Together, these data provide insights into natural variation of histone modifications and their association with gene expression differences between Arabidopsis ecotypes.

Structural and Functional Analyses of Hub Micro RNAs in An Integrated Gene Regulatory Network of Arabidopsis

MicroRNAs(miRNAs)are trans-acting small regulatory RNAs that work coordinately with transcription factors(TFs)to shape the repertoire of cellular m RNAs available for translation.Despite our growing knowledge of individual plant miRNAs,their global roles in gene regulatory networks remain mostly unassessed.Based on interactions obtained from public databases and curated from the literature,we reconstructed an integrated miRNA network in Arabidopsis that includes 66 core TFs,318 miRNAs,and 1712 downstream genes.We found that miRNAs occupy distinct niches and enrich miRNA-containing feed-forward loops(FFLs),particularly those with miRNAs as intermediate nodes.Further analyses revealed that miRNA-containing FFLs coordinate TFs located in different hierarchical layers and that intertwined miRNA-containing FFLs are associated with party and date miRNA hubs.Using the date hub MIR858A as an example,we performed detailed molecular and genetic analyses of three interconnected miRNA-containing FFLs.These analyses revealed individual functions of the selected miRNA-containing FFLs and elucidated how the date hub miRNA fulfills multiple regulatory roles.Collectively,our findings highlight the prevalence and importance of miRNA-containing FFLs,and provide new insights into the design principles and control logics of miRNA regulatory networks governing gene expression programs in plants.

From molecular basics to agronomic benefits:Insights into noncoding RNA-mediated gene regulation in plants

The development of plants is largely dependent on their growth environment.To better adapt to a particular habitat,plants have evolved various subtle regulatory mechanisms for altering gene expression.Noncoding RNAs(ncRNAs)constitute a major portion of the transcriptomes of eukaryotes.Various ncRNAs have been recognized as important regulators of the expression of genes involved in essential biological processes throughout the whole life cycles of plants.In this review,we summarize the current understanding of the biogenesis and contributions of small nucleolar RNA(snoRNA)-and regulatory long noncoding RNA(lncRNA)-mediated gene regulation in plant development and environmental responses.Many regulatory ncRNAs appear to be associated with increased yield,quality and disease resistance of various species and cultivars.These ncRNAs may potentially be used as genetic resources for improving agronomic traits and for molecular breeding.The challenges in understanding plant ncRNA biology and the possibilities to make better use of these valuable gene resources in the future are discussed in this review.

Arabidopsis noncoding RNA modulates seedling greening during deetiolation

Seedling greening is essential for the survival of plants emerging from the soil. The abundance of chlorophyll precursors, including protochlorophyllide(Pchlide), is precisely controlled during the dark-to-light transition, as over-accumulation of Pchlide can lead to cellular photooxidative damage. Previous studies have identified and characterized multiple regulators controlling this important process. HID1(hidden treasure 1) is the first noncoding RNA(ncRNA) found in photomorphogenesis. Under continuous red light,HID1 has been shown to inhibit hypocotyl elongation by repressing the transcription of PIF3(phytochrome interacting factor 3).Here, we report that HID1 acts as a negative regulator of cotyledon greening. Knockdown of HID1 resulted in an increased greening rate of etiolated seedlings relative to wild type when exposed to white light. Genetically, HID1 acts downstream of PIF3 during the dark-to-light transition. The expression of HID1 is not regulated by PIF3 in the dark. Molecularly, the Pchlide content was reduced in dark-grown hid1 mutants than WT. Meanwhile, transcript levels of the protochlorophyllide oxidoreductases known to catalyze Pchlide to chlorophyllide conversion were significantly increased in hid1 seedlings. Thus, our study reveals an additional role of HID1 in the dark-to-light transition in Arabidopsis. Moreover, these results suggest HID1 could regulate distinct targets in different light-mediated developmental processes, and thus is essential to the control of these mechanisms.