Similar Datasets
Project description:RNA helicases are critical regulators of gene expression and therefore strict regulation of their activities is essential to limit promiscuity and ensure target specify. While some RNA helicases appear dedicated for particular substrates others, such as DHX15, are multifunctional, contributing to more than one gene expression process. Protein cofactors play key roles both in modulating RNA helicase activities and also directing them to appropriate substrate RNAs. The largest family of RNA helicase cofactors, with more than 20 members expressed in human cells, are the G-patch proteins, characterised by a glycine-rich domain via which they associate with specific DEAH/RHA helicases. However, the cognate RNA helicases and cellular functions of numerous human G-patch proteins still remain elusive. Here, we reveal that the G-patch protein GPATCH4 is a cofactor of DHX15 that stimulates its ATPase activity. We show that GPATCH4 interacts with DHX15 in the nucleolus via residues in its G-patch domain and co-migrates with pre-ribosomal particles, wherein loss of DHX15 reduces pre-ribosomal co-migration of GPATCH4. Using a combination of high-throughput sequencing approaches (ChIP, CRAC and small RNA-seq), we discovered that GPATCH4 crosslinks to the ribosomal DNA locus and precursor ribosomal RNAs as well as binding to small nucleolar- and small Cajal body-associated- RNAs that guide rRNA and snRNA modifications. In line with this, using RiboMeth-seq analysis to globally analyse rRNA and snRNA 2’-O-methylation revealed that knockout of GPATCH4 impairs methylation at various sites. Utilizing targeted 2’-O-methylation-sensitive RNase H-based cleavage assays, we demonstrated that the regulation of 2’-O-methylation by GPATCH4 is both dependent on, and independent of, its interaction with DHX15. Intriguingly, the ATPase activity of DHX15 is necessary for efficient methylation of DHX15-dependent sites, suggesting a function of DHX15 in regulating snoRNA-guided 2’-O-methylation of rRNA that requires activation by GPATCH4. Overall, our findings not only extend knowledge on RNA helicase regulation by G-patch proteins but also provide important new insights into how the installation of rRNA and snRNA modifications, which are essential for ribosome assembly/function and pre-mRNA splicing, is regulated.
2024-01-30 | GSE230431 | GEO
Project description:Strict regulation of RNA helicase activity, often accomplished by protein cofactors, is essential to ensure target specify within the complex cellular environment. The largest family of RNA helicase cofactors are the G-patch proteins, but the cognate RNA helicases and cellular functions of numerous human G-patch proteins remain elusive. Here, we discover that GPATCH4 is a stimulatory cofactor of DHX15 that interacts with the helicase in the nucleolus via residues in its G-patch domain. We reveal that GPATCH4 associates with pre-ribosomal particles, and crosslinks to the transcribed ribosomal DNA locus and precursor ribosomal RNAs as well as binding to small nucleolar- and small Cajal body-associated- RNAs that guide rRNA and snRNA modifications. Global analysis of rRNA and snRNA 2’-O-methylation revealed that lack of GPATCH4 impairs methylation at various sites. We further demonstrated that the regulation of 2’-O-methylation by GPATCH4 is both dependent on, and independent of, its interaction with DHX15. Intriguingly, the ATPase activity of DHX15 is necessary for efficient methylation of DHX15-dependent sites, suggesting a function of DHX15 in regulating snoRNA-guided 2’-O-methylation of rRNA that requires activation by GPATCH4. Overall, our findings extend knowledge on RNA helicase regulation by G-patch proteins and also provide important new insights into the mechanisms regulating installation of rRNA and snRNA modifications, which are essential for ribosome function and pre-mRNA splicing.
2024-01-30 | GSE235949 | GEO
Project description:RNA helicases are critical regulators of gene expression and therefore strict regulation of their activities is essential to limit promiscuity and ensure target specify. While some RNA helicases appear dedicated for particular substrates others, such as DHX15, are multifunctional, contributing to more than one gene expression process. Protein cofactors play key roles both in modulating RNA helicase activities and also directing them to appropriate substrate RNAs. The largest family of RNA helicase cofactors, with more than 20 members expressed in human cells, are the G-patch proteins, characterised by a glycine-rich domain via which they associate with specific DEAH/RHA helicases. However, the cognate RNA helicases and cellular functions of numerous human G-patch proteins still remain elusive. Here, we reveal that the G-patch protein GPATCH4 is a cofactor of DHX15 that stimulates its ATPase activity. We show that GPATCH4 interacts with DHX15 in the nucleolus via residues in its G-patch domain and co-migrates with pre-ribosomal particles, wherein loss of DHX15 reduces pre-ribosomal co-migration of GPATCH4. Using a combination of high-throughput sequencing approaches (ChIP, CRAC and small RNA-seq), we discovered that GPATCH4 crosslinks to the ribosomal DNA locus and precursor ribosomal RNAs as well as binding to small nucleolar- and small Cajal body-associated- RNAs that guide rRNA and snRNA modifications. In line with this, using RiboMeth-seq analysis to globally analyse rRNA and snRNA 2’-O-methylation revealed that knockout of GPATCH4 impairs methylation at various sites. Utilizing targeted 2’-O-methylation-sensitive RNase H-based cleavage assays, we demonstrated that the regulation of 2’-O-methylation by GPATCH4 is both dependent on, and independent of, its interaction with DHX15. Intriguingly, the ATPase activity of DHX15 is necessary for efficient methylation of DHX15-dependent sites, suggesting a function of DHX15 in regulating snoRNA-guided 2’-O-methylation of rRNA that requires activation by GPATCH4. Overall, our findings not only extend knowledge on RNA helicase regulation by G-patch proteins but also provide important new insights into how the installation of rRNA and snRNA modifications, which are essential for ribosome assembly/function and pre-mRNA splicing, is regulated.
2024-01-30 | GSE230637 | GEO
Project description:RNA helicases are critical regulators of gene expression and therefore strict regulation of their activities is essential to limit promiscuity and ensure target specify. While some RNA helicases appear dedicated for particular substrates others, such as DHX15, are multifunctional, contributing to more than one gene expression process. Protein cofactors play key roles both in modulating RNA helicase activities and also directing them to appropriate substrate RNAs. The largest family of RNA helicase cofactors, with more than 20 members expressed in human cells, are the G-patch proteins, characterised by a glycine-rich domain via which they associate with specific DEAH/RHA helicases. However, the cognate RNA helicases and cellular functions of numerous human G-patch proteins still remain elusive. Here, we reveal that the G-patch protein GPATCH4 is a cofactor of DHX15 that stimulates its ATPase activity. We show that GPATCH4 interacts with DHX15 in the nucleolus via residues in its G-patch domain and co-migrates with pre-ribosomal particles, wherein loss of DHX15 reduces pre-ribosomal co-migration of GPATCH4. Using a combination of high-throughput sequencing approaches (ChIP, CRAC and small RNA-seq), we discovered that GPATCH4 crosslinks to the ribosomal DNA locus and precursor ribosomal RNAs as well as binding to small nucleolar- and small Cajal body-associated- RNAs that guide rRNA and snRNA modifications. In line with this, using RiboMeth-seq analysis to globally analyse rRNA and snRNA 2’-O-methylation revealed that knockout of GPATCH4 impairs methylation at various sites. Utilizing targeted 2’-O-methylation-sensitive RNase H-based cleavage assays, we demonstrated that the regulation of 2’-O-methylation by GPATCH4 is both dependent on, and independent of, its interaction with DHX15. Intriguingly, the ATPase activity of DHX15 is necessary for efficient methylation of DHX15-dependent sites, suggesting a function of DHX15 in regulating snoRNA-guided 2’-O-methylation of rRNA that requires activation by GPATCH4. Overall, our findings not only extend knowledge on RNA helicase regulation by G-patch proteins but also provide important new insights into how the installation of rRNA and snRNA modifications, which are essential for ribosome assembly/function and pre-mRNA splicing, is regulated.
2024-01-30 | GSE230432 | GEO
Project description:Many aspects of membrane-trafficking events are regulated by Rab-family small GTPases. Neurite outgrowth requires massive addition of proteins and lipids to the tips of growing neurites by membrane trafficking, and although several Rabs, including Rab8, Rab10, and Rab11, have been implicated in this process, their regulatory mechanisms during neurite outgrowth are poorly understood. Here, we show that Rabin8, a Rab8-guanine nucleotide exchange factor (GEF), regulates nerve growth factor (NGF)-induced neurite outgrowth of PC12 cells. Knockdown of Rabin8 results in inhibition of neurite outgrowth, whereas overexpression promotes it. We also find that Rab10 is a novel substrate of Rabin8 and that both Rab8 and Rab10 function during neurite outgrowth downstream of Rabin8. Surprisingly, however, a GEF activity-deficient isoform of Rabin8 also promotes neurite outgrowth, indicating the existence of a GEF activity-independent role of Rabin8. The Arf6/Rab8-positive recycling endosomes (Arf6/Rab8-REs) and Rab10/Rab11-positive REs (Rab10/Rab11-REs) in NGF-stimulated PC12 cells are differently distributed. Rabin8 localizes on both RE populations and appears to activate Rab8 and Rab10 there. These localizations and functions of Rabin8 are Rab11 dependent. Thus Rabin8 regulates neurite outgrowth both by coordinating with Rab8, Rab10, and Rab11 and by a GEF activity-independent mechanism.
| S-EPMC4927283 | biostudies-literature
Project description:Regulatory T cells (Tregs) are indispensable for the establishment of tolerance of self-antigens in animals. The transcriptional regulator Foxp3 is critical for Treg development and function, controlling the expression of genes important for Tregs through interactions with binding partners. We previously reported KAP1 as a binding partner of FOXP3 in human Tregs, but the mechanisms by which KAP1 affects Treg function were unclear. In this study, we analyzed mice with Treg-specific deletion of KAP1 and found that they develop spontaneous autoimmune disease. KAP1-deficient Tregs failed to induce Foxp3-regulated Treg signature genes. In addition, KAP1-deficient Tregs were less proliferative due to the decreased expression of Slc1a5, whose expression was KAP1 dependent but Foxp3 independent. This reduced expression of Slc1a5 resulted in reduced mTORC1 activation. Thus, our data suggest that KAP1 regulates Treg function in a Foxp3-dependent manner and also controls Treg proliferation in a Foxp3-independent manner.
| S-EPMC5947873 | biostudies-literature
Project description:The supernumerary chromosome 21 in Down syndrome differentially affects the methylation statuses at CpG dinucleotide sites and creates genome-wide transcriptional dysregulation of parental alleles, ultimately causing diverse pathologies. At present, it is unknown whether those effects are dependent or independent of the parental origin of the nondisjoined chromosome 21. Linkage analysis is a standard method for the determination of the parental origin of this aneuploidy, although it is inadequate in cases with deficiency of samples from the progenitors. Here, we assessed the reliability of the epigenetic 5mCpG imprints resulting in the maternally (oocyte)-derived allele methylation at a differentially methylated region (DMR) of the candidate imprinted WRB gene for asserting the parental origin of chromosome 21. We developed a methylation-sensitive restriction enzyme-specific PCR assay, based on the WRB DMR, across single nucleotide polymorphisms (SNPs) to examine the methylation statuses in the parental alleles. In genomic DNA from blood cells of either disomic or trisomic subjects, the maternal alleles were consistently methylated, while the paternal alleles were unmethylated. However, the supernumerary chromosome 21 did alter the methylation patterns at the RUNX1 (chromosome 21) and TMEM131 (chromosome 2) CpG sites in a parent-of-origin-independent manner. To evaluate the 5mCpG imprints, we conducted a computational comparative epigenomic analysis of transcriptome RNA sequencing (RNA-Seq) and histone modification expression patterns. We found allele fractions consistent with the transcriptional biallelic expression of WRB and ten neighboring genes, despite the similarities in the confluence of both a 17-histone modification activation backbone module and a 5-histone modification repressive module between the WRB DMR and the DMRs of six imprinted genes. We concluded that the maternally inherited 5mCpG imprints at the WRB DMR are uncoupled from the parental allele expression of WRB and ten neighboring genes in several tissues and that trisomy 21 alters DNA methylation in parent-of-origin-dependent and -independent manners.
| S-EPMC4839675 | biostudies-literature
Project description:Fine-tuning of osteoclast differentiation (OD) and bone remodeling is crucial for bone homeostasis. Dissecting the mechanisms regulating osteoclastogenesis is fundamental to understanding the pathogenesis of various bone disorders including osteoporosis and arthritis. Nuclear factor erythroid 2-related factor 1 (NFE2L1, also known as NRF1), which belongs to the CNC-bZIP family of transcription factors, orchestrates a variety of physiological processes and stress responses. While Nfe2l1 gene may be transcribed into multiple alternatively spliced isoforms, the biological function of the different isoforms of NFE2L1 in bone metabolism, osteoclastogenesis in particular, has not been reported. Here we demonstrate that knockout of all isoforms of Nfe2l1 transcripts specifically in the myeloid lineage in mice [Nfe2l1(M)-KO] results in increased activity of osteoclasts, decreased bone mass and worsening of osteoporosis induced by ovariectomy and aging. In comparison, LysM-Cre-mediated Nfe2l1 deletion has no significant effect on the osteoblast and osteocytes. Mechanistic investigations using bone marrow cells and RAW 264.7 cells revealed that deficiency of Nfe2l1 leads to accelerated and elevated OD, which is attributed, at least in part, to enhanced accumulation of ROS in the early stage of OD and expression of nuclear factor of activated T cells, cytoplasmic, calcineurin dependent 1α (Nfatc1/α). In addition, NFE2L1 regulates the transcription of multiple antioxidant genes and Nfatc1/α and OD in an isoform-specific manner. While long isoforms of NFE2L1 function as accelerators of induction of Nfatc1/α and antioxidant genes and OD, the short isoform NFE2L1-453 serves as a brake that keeps the long isoforms' accelerator effects in check. These findings provide a novel insight into the regulatory roles of NFE2L1 in osteoclastogenesis and highlight that NFE2L1 is essential in regulating bone remodeling and thus may be a valuable therapeutic target for bone disorders.
| S-EPMC8591424 | biostudies-literature
Project description:Although tetrabromobisphenol A (TBBPA) has been well proven to disturb TH signaling in both in vitro and in vivo assays, it is still unclear whether TBBPA can affect brain development due to TH signaling disruption. Here, we employed the T3-induced Xenopus metamorphosis assay (TIXMA) and the spontaneous metamorphosis assay to address this issue. In the TIXMA, 5-500 nmol/L TBBPA affected T3-induced TH-response gene expression and T3-induced brain development (brain morphological changes, cell proliferation, and neurodifferentiation) at premetamorphic stages in a complicated biphasic concentration-response manner. Notably, 500 nmol/L TBBPA treatment alone exerted a stimulatory effect on tadpole growth and brain development at these stages, in parallel with a lack of TH signaling activation, suggesting the involvement of other signaling pathways. As expected, at the metamorphic climax, we observed inhibitory effects of 50-500 nmol/L TBBPA on metamorphic development and brain development, which was in agreement with the antagonistic effects of higher concentrations on T3-induced brain development at premetamorphic stages. Taken together, all results demonstrate that TBBPA can disturb TH signaling and subsequently interfere with TH-dependent brain development in Xenopus; meanwhile, other signaling pathways besides TH signaling could be involved in this process. Our study improves the understanding of the effects of TBBPA on vertebrate brain development.
| S-EPMC8746619 | biostudies-literature
Project description:CMTR1 contributes to mRNA cap formation by methylating the first transcribed nucleotide ribose at the O-2 position. mRNA cap O-2 methylation has roles in mRNA stabilisation and translation, and self-RNA tolerance in innate immunity. We report that CMTR1 is recruited to serine-5-phosphorylated RNA Pol II C-terminal domain, early in transcription. We isolated CMTR1 in a complex with DHX15, an RNA helicase functioning in splicing and ribosome biogenesis, and characterised it as a regulator of CMTR1. When DHX15 is bound, CMTR1 activity is repressed and the methyl-transferase does not bind to RNA pol II. Conversely, CMTR1 activates DHX15 helicase activity, which is likely to impact several nuclear functions. In HCC1806 breast carcinoma cell line, the DHX15-CMTR1 interaction controls ribosome loading of a subset of mRNAs and regulates cell proliferation. The impact of the CMTR1-DHX15 interaction is complex and will depend on the relative expression of these enzymes and their interactors, and the cellular dependency on different RNA processing pathways.
| S-EPMC6071836 | biostudies-literature