Inputs and outputs for chromatin-targeted RNAi

Plant gene silencing is targeted to transposons and repeated sequences by small RNAs from the RNA interference (RNAi) pathway. Like classical RNAi, RNA-directed chromatin silencing involves the cleavage of double-stranded RNA by Dicer endonucleases to create small interfering RNAs (siRNAs), which bind to the Argonaute protein. The production of double-stranded RNA (dsRNA) must be carefully controlled to prevent inappropriate silencing. A plant-specific RNA polymerase IV (Pol IV) initiates siRNA production at silent heterochromatin, but Pol IV-independent mechanisms for making dsRNA also exist. Downstream of siRNA biogenesis, multiple chromatin marks might be targeted by Argonaute-siRNA complexes, yet mechanisms of chromatin modification remain poorly understood. Genomic studies of siRNA target loci promise to reveal novel biological functions for chromatin-targeted RNAi.     

RNA干扰与染色质沉默——生物体内精密的网络调控机制

基因表达受不同层次的调控.RNA干扰通过产生双链小RNA诱导同源mRNA序列降解,从而在转录后抑制特定基因的表达.最新的研究成果显示：RNA干扰产生的双链小RNA可通过与染色质中的重复序列DNA及组蛋白甲基化酶相互作用,引起组蛋白H3 Lys9的甲基化,进一步与异染色质形成相关蛋白结合,导致染色质沉默.综述了RNA干扰,小RNA,组蛋白修饰,染色质沉默及基因表达调控之间存在着精密的网络调控机制.     

Identification of noncoding transcripts from within CENP-A chromatin at fission yeast centromeres

The histone H3 variant CENP-A is the most favored candidate for an epigenetic mark that specifies the centromere. In fission yeast, adjacent heterochromatin can direct CENP-A(Cnp1) chromatin establishment, but the underlying features governing where CENP-A(Cnp1) chromatin assembles are unknown. We show that, in addition to centromeric regions, a low level of CENP-A(Cnp1) associates with gene promoters where histone H3 is depleted by the activity of the Hrp1(Chd1) chromatin-remodeling factor. Moreover, we demonstrate that noncoding RNAs are transcribed by RNA polymerase II (RNAPII) from CENP-A(Cnp1) chromatin at centromeres. These analyses reveal a similarity between centromeres and a subset of RNAPII genes and suggest a role for remodeling at RNAPII promoters within centromeres that influences the replacement of histone H3 with CENP-A(Cnp1).     

The small RNA profile during Drosophila melanogaster development

Small RNAs ranging in size between 20 and 30 nucleotides are involved in different types of regulation of gene expression including mRNA degradation, translational repression, and chromatin modification. Here we describe the small RNA profile of Drosophila melanogaster as a function of development. We have cloned and sequenced over 4000 small RNAs, 560 of which have the characteristics of RNase III cleavage products. A nonredundant set of 62 miRNAs was identified. We also isolated 178 repeat-associated small interfering RNAs (rasiRNAs), which are cognate to transposable elements, satellite and microsatellite DNA, and Suppressor of Stellate repeats, suggesting that small RNAs participate in defining chromatin structure. rasiRNAs are most abundant in testes and early embryos, where regulation of transposon activity is critical and dramatic changes in heterochromatin structure occur.     

The mechanism selecting the guide strand from small RNA duplexes is different among argonaute proteins

Double-stranded RNA induces RNA silencing and is cleaved into21–24 nt small RNA duplexes by Dicer enzyme. A strandof Dicer-generated small RNA duplex (called the guide strand)is then selected by a thermodynamic mechanism to associate withArgonaute (AGO) protein. This AGO–small RNA complex functionsto cleave mRNA, repress translation or modify chromatin structurein a sequence-specific manner. Although a model plant, Arabidopsisthaliana, contains 10 AGO genes, their roles and molecular mechanismsremain obscure. In this study, we analyzed the roles of ArabidopsisAGO2 and AGO5. Interestingly, the 5' nucleotide of small RNAsthat associated with AGO2 was mainly adenine (85.7%) and thatwith AGO5 was mainly cytosine (83.5%). Small RNAs that wereabundantly cloned from the AGO2 immunoprecipitation fraction(miR163-LL, which is derived from the Lower Left of mature miR163in pre-miR163, and miR390) and from the AGO5 immunoprecipitationfraction (miR163-UL, which is derived from the Upper Left ofmature miR163 in pre-miR163, and miR390^*) are derived from thesingle small RNA duplexes, miR163-LL/miR163-UL and miR390/miR390^*.Each strand of the miR163-LL/miR163-UL duplex is selectivelysorted to associate with AGO2 or AGO5 in a 5' nucleotide-dependentmanner rather than in a thermodynamic stability-dependent manner.Furthermore, we showed that both AGO2 and AGO5 have the abilityto bind cucumber mosaic virus-derived small RNAs. These resultsclearly indicate that the mechanism selecting the guide strandis different among AGO proteins and that multiple AGO genesare involved in anti-virus defense in plants.