Project description:Short interspersed nuclear elements (SINEs) are retrotransposons evolutionarily derived from endogenous RNA Polymerase III RNAs. Though SINE elements have undergone exaptation into gene regulatory elements, how transcribed SINE RNA impacts transcriptional and post-transcriptional regulation is largely unknown. This is partly due to a lack of information regarding which of the loci have transcriptional potential. Here, we present an approach (short interspersed nuclear element sequencing, SINE-seq), which selectively profiles RNA Polymerase III-derived SINE RNA, thereby identifying transcriptionally active SINE loci. Applying SINE-seq to monitor murine B2 SINE expression during a gammaherpesvirus infection revealed transcription from 28,270 SINE loci, with ~50% of active SINE elements residing within annotated RNA Polymerase II loci. Furthermore, B2 RNA can form intermolecular RNA-RNA interactions with complementary mRNAs, leading to nuclear retention of the targeted mRNA via a mechanism involving p54nrb. These findings illuminate a pathway for the selective regulation of mRNA export during stress via retrotransposon activation.
Project description:Short interspersed element (SINE) RNAs are upregulated by cellular stresses1 (heat shock, DNA damage and viral infection) and accumulate in human diseases (macular degeneration2, lupus3, and Alzheimer’s disease4). These transcripts activate inflammasomes5, a family of cytoplasmic multiprotein complexes that sense danger molecules and initiate innate immune responses by activating caspase-1-dependent cytokine production and inflammatory death6, but the molecular sensor of SINE RNAs is unknown. Here, we identify DDX17, a member of the DEAD box family of RNA helicases7, as a sensor of SINE RNAs requisite for inflammasome activation. Induction of caspase-1 cleavage and release of IL-1 and IL-18 by SINE RNAs requires dual recruitment of NLRP3 and NLRC4 but proceeds independent of NAIPs, immune sensors required for canonical NLRC4 activation by bacterial proteins8,9. Instead, SINE RNAs trigger DDX17–NLRC4 interaction, which licenses inflammasome activation. We also report increased levels of DDX17 protein and association of DDX17 and NLRC4 in the retinal pigmented epithelium (RPE) of human eyes with an advanced, untreatable form of age-related macular degeneration (AMD). Disrupting DDX17–NLRC4 signalling blocks SINE RNA-induced inflammasome activation in human RPE cells and RPE degeneration in an animal model of AMD. Our findings uncover a non-canonical mode of inflammasome activation by endogenous retrotransposon transcripts, and provide new potential targets for macular degeneration and potentially other diseases.
Project description:Plants monitor and integrate temperature, photoperiod and light quality signals to respond to continuous changes in their environment. The GIGANTEA (GI) protein is central in diverse signaling pathways, including photoperiodic, sugar and light signaling pathways, stress responses and circadian clock regulation. Previously, GI was shown to activate expression of the key floral regulators CONSTANS (CO) and FLOWERING LOCUS T (FT) by facilitating degradation of a family of CYCLING DOF FACTOR (CDF) transcriptional repressors. However, whether CDFs are implicated in other processes regulated by GI remains unclear. We investigated the contribution of the GI-CDF module to traits that depend on GI. Transcriptome profiling indicated that mutations in GI and the CDFs have antagonistic effects on expression of a wider set of genes than CO and FT, whilst other genes are regulated by GI independently of the CDFs. Detailed expression studies followed by phenotypic assays showed that the CDFs function downstream of GI to control responses to freezing temperatures and growth, but are not necessary for proper clock function. Thus GI-mediated regulation of CDFs contributes to several processes in addition to flowering, but is not implicated in all of the phenotypes influenced by GI. Seedlings of Col-0, the cdf1-R cdf2-1 cdf3-1 cdf5-1 quadruple mutant, gi-100 and gi-100 cdf1-R cdf2-1 cdf3-1 cdf5-1 quintuple mutant were grown for 10 days under LD conditions (16h light/8h dark). The aerial part of the seedlings was collected at ZT12 and RNA was prepared from three biological replicas
Project description:Plants monitor and integrate temperature, photoperiod and light quality signals to respond to continuous changes in their environment. The GIGANTEA (GI) protein is central in diverse signaling pathways, including photoperiodic, sugar and light signaling pathways, stress responses and circadian clock regulation. Previously, GI was shown to activate expression of the key floral regulators CONSTANS (CO) and FLOWERING LOCUS T (FT) by facilitating degradation of a family of CYCLING DOF FACTOR (CDF) transcriptional repressors. However, whether CDFs are implicated in other processes regulated by GI remains unclear. We investigated the contribution of the GI-CDF module to traits that depend on GI. Transcriptome profiling indicated that mutations in GI and the CDFs have antagonistic effects on expression of a wider set of genes than CO and FT, whilst other genes are regulated by GI independently of the CDFs. Detailed expression studies followed by phenotypic assays showed that the CDFs function downstream of GI to control responses to freezing temperatures and growth, but are not necessary for proper clock function. Thus GI-mediated regulation of CDFs contributes to several processes in addition to flowering, but is not implicated in all of the phenotypes influenced by GI.
Project description:We identified a novel long non-coding RNA Lx8-SINE B2, that is a marker of pluripotency. Depletion of Lx8-SINE B2 impacts embryonic stem cell self-renewal. RNA-seq analysis of Lx8-SINE B2 depletion revealed that a number of glycolytic genes with decreased expression. Mechanistically, we found that the Lx8-SINE B2 activates the glycolysis pathway by binding to Eno1. Collectively, our data suggest that Lx8-SINE B2 maintains the self-renewal of mESCs through glycolysis.
Project description:H3K27ac HiChIP analysis was performed in GI-MEN DOX-ASCL1 cells to analyze active chromatin-chromatin interactions in GI-MEN DOX-ASCL1 cells.
Project description:Cut & Run analysis was performed in an neuroblastoma cell line to analyze DNA bindings of ASCL1-tag-HA in GI-MEN ASCL1-tag-HA cells and GI-MEN ASCL1-tag-HA+4TFs cells; analyze DNA bindings of MYCN, PHOX2B and H3K27ac in, GI-MEN 4TFs cells, and GI-MEN ASCL1-tag-HA+4TFs cells.