Project description:MicroRNAs (miRNAs) and small-interfering RNAs (siRNAs) negatively regulate their targets by 1) repressing translation, 2) endonucleolytic RNA cleavage, or 3) DNA methylation resulting in transcriptional silencing. P-body/decapping components are likely required for translational repression, but are not known to function in other posttranscriptional regulatory pathways or to affect smRNA levels. Here, we show that the P-body/decapping protein DCP5 is required for miRNA-mediated translational repression but not cleavage, and to regulate the transcription of specific miRNAs. We find that this protein also affects the abundance of tRNA-derived smRNAs. Significantly, DCP5 is required for the transcriptional silencing and DNA methylation of numerous transposable/repetitive elements and imprinted genes, indicating that it is a novel component of the RNA-directed DNA methylation pathway. Our results demonstrate that DCP5 and likely the P-body itself are required for multiple smRNA-mediated silencing pathways and provide the first evidence for the spatial separation of translational inhibition and cleavage by miRNAs. small RNA (smRNA) expression comparison between wildtype (Col-0) and dcp5 mutant plants in Arabidopsis
Project description:MicroRNAs (miRNAs) and small-interfering RNAs (siRNAs) negatively regulate their targets by 1) repressing translation, 2) endonucleolytic RNA cleavage, or 3) DNA methylation resulting in transcriptional silencing. P-body/decapping components are likely required for translational repression, but are not known to function in other posttranscriptional regulatory pathways or to affect smRNA levels. Here, we show that the P-body/decapping protein DCP5 is required for miRNA-mediated translational repression but not cleavage, and to regulate the transcription of specific miRNAs. We find that this protein also affects the abundance of tRNA-derived smRNAs. Significantly, DCP5 is required for the transcriptional silencing and DNA methylation of numerous transposable/repetitive elements and imprinted genes, indicating that it is a novel component of the RNA-directed DNA methylation pathway. Our results demonstrate that DCP5 and likely the P-body itself are required for multiple smRNA-mediated silencing pathways and provide the first evidence for the spatial separation of translational inhibition and cleavage by miRNAs. total RNA expression comparison with between wildtype (Col-0) and dcp5 mutant plants in Arabidopsis
Project description:MicroRNAs (miRNAs) and small-interfering RNAs (siRNAs) negatively regulate their targets by 1) repressing translation, 2) endonucleolytic RNA cleavage, or 3) DNA methylation resulting in transcriptional silencing. P-body/decapping components are likely required for translational repression, but are not known to function in other posttranscriptional regulatory pathways or to affect smRNA levels. Here, we show that the P-body/decapping protein DCP5 is required for miRNA-mediated translational repression but not cleavage, and to regulate the transcription of specific miRNAs. We find that this protein also affects the abundance of tRNA-derived smRNAs. Significantly, DCP5 is required for the transcriptional silencing and DNA methylation of numerous transposable/repetitive elements and imprinted genes, indicating that it is a novel component of the RNA-directed DNA methylation pathway. Our results demonstrate that DCP5 and likely the P-body itself are required for multiple smRNA-mediated silencing pathways and provide the first evidence for the spatial separation of translational inhibition and cleavage by miRNAs.
Project description:MicroRNAs (miRNAs) and small-interfering RNAs (siRNAs) negatively regulate their targets by 1) repressing translation, 2) endonucleolytic RNA cleavage, or 3) DNA methylation resulting in transcriptional silencing. P-body/decapping components are likely required for translational repression, but are not known to function in other posttranscriptional regulatory pathways or to affect smRNA levels. Here, we show that the P-body/decapping protein DCP5 is required for miRNA-mediated translational repression but not cleavage, and to regulate the transcription of specific miRNAs. We find that this protein also affects the abundance of tRNA-derived smRNAs. Significantly, DCP5 is required for the transcriptional silencing and DNA methylation of numerous transposable/repetitive elements and imprinted genes, indicating that it is a novel component of the RNA-directed DNA methylation pathway. Our results demonstrate that DCP5 and likely the P-body itself are required for multiple smRNA-mediated silencing pathways and provide the first evidence for the spatial separation of translational inhibition and cleavage by miRNAs.
Project description:Arabidopsis DCP5, a homolog of human RNA-associated protein 55, is a nessary component of eukaryotic processing bodies (P-bodies). knockdown mutant of dcp5-1 showed compromised RNA decapping activity and reduced P-body size. Here we profiled Arabidopsis transcriptome of roots, shoots, and inflorensences in Col-0 and DCP5-1 mutant using strand-specific RNA-sequencing. Our analysis identified a large number of DCP5-regulatd transcripts in Arabidopsis.
Project description:Epigenetic mechanisms oversee epidermal homeostasis and oncogenesis. The identification of kinases controlling these processes has direct therapeutic implications. We show that ULK3 is a nuclear kinase with elevated expression levels in squamous cell carcinomas (SCCs) arising in multiple body sites, including skin and Head/Neck. ULK3 loss by gene silencing or deletion reduces proliferation and clonogenicity of human keratinocytes and SCC-derived cells and affects transcription impinging on stem cell-related and metabolism programs. Mechanistically, ULK3 directly binds and regulates the activity of two histone arginine methyltransferases, PRMT1 and PRMT5 (PRMT1/5), with ULK3 loss compromising PRMT1/5 chromatin association to specific genes and overall methylation of histone H4, a shared target of these enzymes. These findings are of translational significance, as downmodulating ULK3 by RNA interference or locked antisense nucleic acids (LNAs) blunts the proliferation and tumorigenic potential of SCC cells and promotes differentiation in two orthotopic models of skin cancer.
Project description:Polycomb Repressive Complexes 1 and 2 (PRC1 and PRC2) are histone-modifying and -binding complexes that mediate the formation of facultative heterochromatin and are required for silencing of developmental genes and maintenance of cell fate. Multiple pathways of RNA decay work together to establish and maintain heterochromatin in fission yeast, including a recently identified role for a conserved RNA degradation complex called the rixosome or RIX1 complex. Whether RNA degradation also plays a role in the stability of mammalian heterochromatin remains unknown. Here we show that the rixosome contributes to silencing of many Polycomb targets in human cells. The rixosome associates with human PRC complexes and is enriched at promoters of Polycomb target genes. Importantly, depletion of either the rixosome or Polycomb results in accumulation of paused and elongating RNA polymerase at Polycomb-target genes. We identify point mutations in the RING1B subunit of PRC1 that disrupt the interaction between PRC1 and the rixosome and result in diminished silencing, suggesting that direct recruitment of the rixosome to chromatin is required for silencing. Finally, we show that the RNA kinase activity of the rixosome and the XRN2 exoribonuclease, which degrades RNAs with 5’ mono-phosphate groups generated by the rixosome, are required for silencing. Our findings suggest that rixosome-mediated degradation of nascent RNA is conserved from fission yeast to human, although in human cells the rixosome degrades RNA in facultative rather than constitutive heterochromatin.