Profiling transcriptome composition and dynamics within nuclear compartments using SLAM-RT&Tag
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE272219 | GEO | 2025/03/04
REPOSITORIES: GEO
Similar Datasets
Project description:Nuclear speckles, a type of membraneless nuclear organelle in higher eukaryotic cells, play a vital role in gene expression regulation. Using the reverse transcription-based RBP binding sites sequencing (ARTR-seq) method, we study human transcripts associated with nuclear speckles. We identify three gene groups whose transcripts demonstrate different speckle localization properties and dynamics – stably enriched in nuclear speckle post-transcriptionally, transiently enriched in speckles at the pre-mRNA stage co-transcriptionally, and not enriched in speckles. We show that nuclear speckles specifically facilitate splicing of speckle-enriched transcripts. We further reveal RNA sequence features contributing to transcript speckle localization, underscoring a tight interplay between genome organization, RNA cis-elements, and transcript speckle enrichment, and connecting transcript speckle localization with splicing logic. Finally, we show that speckles can act as hubs for the regulated retention of introns during cellular stress. Collectively, our data highlight a role of nuclear speckles in both co- and post-transcriptional splicing regulation.
2024-09-05 | GSE255207 | GEO
Project description:Nuclear speckles, a type of membraneless nuclear organelle in higher eukaryotic cells, play a vital role in gene expression regulation. Using the reverse transcription-based RBP binding sites sequencing (ARTR-seq) method, we study human transcripts associated with nuclear speckles. We identify three gene groups whose transcripts demonstrate different speckle localization properties and dynamics – stably enriched in nuclear speckle post-transcriptionally, transiently enriched in speckles at the pre-mRNA stage co-transcriptionally, and not enriched in speckles. We show that nuclear speckles specifically facilitate splicing of speckle-enriched transcripts. We further reveal RNA sequence features contributing to transcript speckle localization, underscoring a tight interplay between genome organization, RNA cis-elements, and transcript speckle enrichment, and connecting transcript speckle localization with splicing logic. Finally, we show that speckles can act as hubs for the regulated retention of introns during cellular stress. Collectively, our data highlight a role of nuclear speckles in both co- and post-transcriptional splicing regulation.
2024-09-05 | GSE255206 | GEO
Project description:Prior work revealed nuclear-localized aminoacyl-tRNA synthetases (aaRSs) that checked newly synthesized tRNAs for charging before export to the cytoplasm. To reveal other functions, we sought to identify nuclear proteins that interact with arginyl-tRNA synthetase (ArgRS) in the nucleus. Serine/Arginine Repetitive Matrix Protein 2 (SRRM2), which is stored with RNA splicing apparatus components in nuclear speckle condensates, was found as a consistent interaction partner that co localized with SRRM2 ArgRS in nuclear speckles. Dynamic photo-bleaching experiments showed that, consistent with condensate properties, SRRM2 has a fluctuating appearance in speckles. Knock down of ArgRS impeded SRRM2 speckle trafficking and, coincidently, altered splicing processing of pre-mRNA transcripts. Among the altered spliced variants, those of tRNA synthetase family members were prominent. Thus, nuclear ArgRS shapes the dynamics of a protein in class of nuclear condensates. Also, the work expands the repertoire of nuclear tRNA synthetase roles to include regulation of RNA splicing, including of aaRS family member transcripts.
2023-04-14 | PXD015692 | Pride
Project description:Nuclear speckles are membraneless organelles that associate with active transcription sites and participate in post-transcriptional mRNA processing. During the cell cycle, nuclear speckles dissolve following phosphorylation of their protein components. Here, we identify the PP1 family as the phosphatases that counteract kinase-mediated dissolution. PP1 overexpression increases speckle cohesion and leads to retention of polyadenylated RNA within speckles and the nucleus. Using APEX2 proximity labeling combined with RNA-sequencing, we characterized the relationship between the cohesion of nuclear speckles and the recruitment of specific RNAs. We find that many transcripts are preferentially enriched within nuclear speckles compared to the nucleoplasm, particularly chromatin- and nucleus-associated transcripts. While total polyadenylated RNA retention increased with nuclear speckle cohesion, the ratios of most mRNA species to each other were constant, indicating non-selective, or proportional, retention. We then explored whether nuclear speckle cohesion changes in response to environmental perturbations associated with changes in kinase or phosphatase activity. We found that cellular responses to heat shock, oxidative stress, and hypoxia include changes to the cohesion of nuclear speckles and mRNA retention. Our results demonstrate that tuning the material properties of nuclear speckles provides a mechanism for the acute control of mRNA localization.
2024-11-28 | PXD055775 | Pride
Project description:Nuclear speckles are membraneless organelles that associate with active transcription sites and participate in post-transcriptional mRNA processing. During mitosis, nuclear speckles dissolve following phosphorylation of their protein components. Here, we identify PP1 phosphatases as responsible for counteracting kinase-mediated dissolution. Their overexpression increases speckle cohesion and leads to retention of polyadenylated RNA within speckles and the nucleus. By performing APEX2 proximity labeling combined with RNA-sequencing, we characterized the association of specific RNAs with nuclear speckles depending on their cohesion state. We find that many transcripts are preferentially enriched within nuclear speckles compared to the nucleoplasm, particularly chromatin- and nucleus-associated transcripts. While total polyadenylated RNA retention increased with greater nuclear speckle cohesion, the ratios of most mRNA species to each other were constant, indicating non-selective, or proportional, retention. We then explored whether nuclear speckle cohesion changes in response to environmental perturbations associated with changes in kinase or phosphatase activity. We found that cellular responses to heat shock, oxidative stress, and hypoxia include changes to the cohesion of nuclear speckles and mRNA retention. Our results demonstrate that tuning the material properties of nuclear speckles provides a mechanism for the acute control of mRNA localization.
2024-11-10 | GSE240883 | GEO
Project description:Nuclear speckles are membraneless organelles that associate with active transcription sites and participate in post-transcriptional mRNA processing. During mitosis, nuclear speckles dissolve following phosphorylation of their protein components. Here, we identify PP1 phosphatases as responsible for counteracting kinase-mediated dissolution. Their overexpression increases speckle cohesion and leads to retention of polyadenylated RNA within speckles and the nucleus. By performing APEX2 proximity labeling combined with RNA-sequencing, we characterized the association of specific RNAs with nuclear speckles depending on their cohesion state. We find that many transcripts are preferentially enriched within nuclear speckles compared to the nucleoplasm, particularly chromatin- and nucleus-associated transcripts. While total polyadenylated RNA retention increased with greater nuclear speckle cohesion, the ratios of most mRNA species to each other were constant, indicating non-selective, or proportional, retention. We then explored whether nuclear speckle cohesion changes in response to environmental perturbations associated with changes in kinase or phosphatase activity. We found that cellular responses to heat shock, oxidative stress, and hypoxia include changes to the cohesion of nuclear speckles and mRNA retention. Our results demonstrate that tuning the material properties of nuclear speckles provides a mechanism for the acute control of mRNA localization.
2024-11-10 | GSE240891 | GEO
Project description:Nuclear speckles are prominent nuclear bodies that contain a myriad of factors involved in gene expression. The role of nuclear speckles as activating transcriptional compartments is emerging. However, the extent that the association between speckles and DNA is regulatable, and the mechanisms that govern regulated speckle association are currently unclear. Using DNA- and RNA-FISH, we show that speckle association can be mediated by the p53 transcription factor, finding that p53 activation drives speckle association of specific p53 transcriptional targets. Analysis of a key p53 target, p21, revealed an increase in nascent transcripts at speckle-adjacent transcription sites, supporting a role for speckles in amplifying transcriptional output. Importantly, p53-regulated speckle association of p21 did not depend on transcriptional activation, demonstrating that speckle association is not merely a consequence of gene expression. In contrast, speckle association of p21 did require DNA binding functions of p53, providing a mechanism for the specificity by which speckle association is regulated. Beyond p21, a substantial subset of p53 targets have p53-regulated speckle association, while other p53 targets do not, and we find that genomic context is highly deterministic of which target genes have regulated speckle association. These findings reveal a novel means by which transcription factors may control gene expression and provide a mechanism for the specificity of regulated speckle association.
2021-01-04 | GSE154095 | GEO
Project description:Nuclear speckles are dynamic nuclear bodies characterized by high local concentrations of RNA binding proteins and specific non-coding RNAs. Although the contents of speckles suggest multifaceted roles in regulating chromatin dynamics and gene expression, the overarching biological function(s) of nuclear speckles remain enigmatic. In this study, we investigate speckle compositional variation in human cancer, finding two main speckle compositional states based on RNA expression of speckle-resident proteins. One cancer speckle state was more similar to normal adjacent tissues, while the other was dissimilar from normal tissue, and thus considered an aberrant cancer speckle state. We link the aberrant speckle state to altered speckle positioning within the nucleus, to elevation of the TREX RNA export complex, and to worse patient outcomes in clear cell renal cell carcinoma (ccRCC). ccRCC is typified by hyperactivation of the HIF-2a transcription factor, and we demonstrate that HIF-2a drives physical association of a select subset of its target genes with nuclear speckles depending on HIF-2a's two speckle targeting motifs (STMs) defined in this study. STMs are highly enriched among transcription factors, suggesting that DNA-speckle targeting may be a general mechanism of gene regulation and providing a resource of candidate speckle-targeting factors. Via integration of tissue culture functional studies with tumor genomic and imaging analysis, we show that HIF-2a gene regulatory programs are impacted by speckle compositional state and by abrogation of speckle targeting abilities of HIF-2a. These findings suggest that, in ccRCC, a key biological function of nuclear speckles is to modulate expression of a specific subset of HIF-2a-regulated target genes that, in turn, influence patient outcomes. Beyond ccRCC, tumor speckle compositional states broadly correlate with altered functional pathways and expression of speckle-associated gene neighborhoods, exposing a general link between nuclear speckles and gene expression dysregulation in human cancer.
2024-09-03 | GSE248710 | GEO
Project description:Nuclear speckles are dynamic nuclear bodies characterized by high local concentrations of RNA binding proteins and specific non-coding RNAs. Although the contents of speckles suggest multifaceted roles in regulating chromatin dynamics and gene expression, the overarching biological function(s) of nuclear speckles remain enigmatic. In this study, we investigate speckle compositional variation in human cancer, finding two main speckle compositional states based on RNA expression of speckle-resident proteins. One cancer speckle state was more similar to normal adjacent tissues, while the other was dissimilar from normal tissue, and thus considered an aberrant cancer speckle state. We link the aberrant speckle state to altered speckle positioning within the nucleus, to elevation of the TREX RNA export complex, and to worse patient outcomes in clear cell renal cell carcinoma (ccRCC). ccRCC is typified by hyperactivation of the HIF-2a transcription factor, and we demonstrate that HIF-2a drives physical association of a select subset of its target genes with nuclear speckles depending on HIF-2a's two speckle targeting motifs (STMs) defined in this study. STMs are highly enriched among transcription factors, suggesting that DNA-speckle targeting may be a general mechanism of gene regulation and providing a resource of candidate speckle-targeting factors. Via integration of tissue culture functional studies with tumor genomic and imaging analysis, we show that HIF-2a gene regulatory programs are impacted by speckle compositional state and by abrogation of speckle targeting abilities of HIF-2a. These findings suggest that, in ccRCC, a key biological function of nuclear speckles is to modulate expression of a specific subset of HIF-2a-regulated target genes that, in turn, influence patient outcomes. Beyond ccRCC, tumor speckle compositional states broadly correlate with altered functional pathways and expression of speckle-associated gene neighborhoods, exposing a general link between nuclear speckles and gene expression dysregulation in human cancer.
2024-09-03 | GSE248709 | GEO
Project description:Nuclear speckles are dynamic nuclear bodies characterized by high local concentrations of RNA binding proteins and specific non-coding RNAs. Although the contents of speckles suggest multifaceted roles in regulating chromatin dynamics and gene expression, the overarching biological function(s) of nuclear speckles remain enigmatic. In this study, we investigate speckle compositional variation in human cancer, finding two main speckle compositional states based on RNA expression of speckle-resident proteins. One cancer speckle state was more similar to normal adjacent tissues, while the other was dissimilar from normal tissue, and thus considered an aberrant cancer speckle state. We link the aberrant speckle state to altered speckle positioning within the nucleus, to elevation of the TREX RNA export complex, and to worse patient outcomes in clear cell renal cell carcinoma (ccRCC). ccRCC is typified by hyperactivation of the HIF-2a transcription factor, and we demonstrate that HIF-2a drives physical association of a select subset of its target genes with nuclear speckles depending on HIF-2a's two speckle targeting motifs (STMs) defined in this study. STMs are highly enriched among transcription factors, suggesting that DNA-speckle targeting may be a general mechanism of gene regulation and providing a resource of candidate speckle-targeting factors. Via integration of tissue culture functional studies with tumor genomic and imaging analysis, we show that HIF-2a gene regulatory programs are impacted by speckle compositional state and by abrogation of speckle targeting abilities of HIF-2a. These findings suggest that, in ccRCC, a key biological function of nuclear speckles is to modulate expression of a specific subset of HIF-2a-regulated target genes that, in turn, influence patient outcomes. Beyond ccRCC, tumor speckle compositional states broadly correlate with altered functional pathways and expression of speckle-associated gene neighborhoods, exposing a general link between nuclear speckles and gene expression dysregulation in human cancer.
2024-09-03 | GSE248708 | GEO