scholarly journals The Sub-Nuclear Localization of RNA-Binding Proteins in KSHV-Infected Cells

Cells ◽  
2020 ◽  
Vol 9 (9) ◽  
pp. 1958
Author(s):  
Ella Alkalay ◽  
Chen Gam Ze Letova Refael ◽  
Irit Shoval ◽  
Noa Kinor ◽  
Ronit Sarid ◽  
...  
Keyword(s):  
Binding Proteins ◽  
Rna Binding ◽  
Viral Infections ◽  
Rna Binding Proteins ◽  
Nuclear Periphery ◽  
Lytic Cycle ◽  
Splicing Factors ◽  
Nuclear Speckles ◽  
Viral Rnas ◽  
Infected Cells

RNA-binding proteins, particularly splicing factors, localize to sub-nuclear domains termed nuclear speckles. During certain viral infections, as the nucleus fills up with replicating virus compartments, host cell chromatin distribution changes, ending up condensed at the nuclear periphery. In this study we wished to determine the fate of nucleoplasmic RNA-binding proteins and nuclear speckles during the lytic cycle of the Kaposi’s sarcoma associated herpesvirus (KSHV). We found that nuclear speckles became fewer and dramatically larger, localizing at the nuclear periphery, adjacent to the marginalized chromatin. Enlarged nuclear speckles contained splicing factors, whereas other proteins were nucleoplasmically dispersed. Polyadenylated RNA, typically found in nuclear speckles under regular conditions, was also found in foci separated from nuclear speckles in infected cells. Poly(A) foci did not contain lncRNAs known to colocalize with nuclear speckles but contained the poly(A)-binding protein PABPN1. Examination of the localization of spliced viral RNAs revealed that some spliced transcripts could be detected within the nuclear speckles. Since splicing is required for the maturation of certain KSHV transcripts, we suggest that the infected cell does not dismantle nuclear speckles but rearranges their components at the nuclear periphery to possibly serve in splicing and transport of viral RNAs into the cytoplasm.

scholarly journals Targeted Quantification of Detergent-Insoluble RNA-Binding Proteins in Human Brain Reveals Stage and Disease Specific Co-aggregation in Alzheimer’s Disease

2021 ◽  
Vol 14 ◽  
Author(s):  
Qi Guo ◽  
Eric B. Dammer ◽  
Maotian Zhou ◽  
Sean R. Kundinger ◽  
Marla Gearing ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Rna Splicing ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Splicing Factors ◽  
Nuclear Speckles ◽  
Spliceosome Assembly ◽  
Network Analyses

Core spliceosome and related RNA-binding proteins aggregate in Alzheimer’s disease (AD) brain even in early asymptomatic stages (AsymAD) of disease. To assess the specificity of RNA-binding protein aggregation in AD, we developed a targeted mass spectrometry approach to quantify broad classes of RNA-binding proteins with other pathological proteins including tau and amyloid beta (Aβ) in detergent insoluble fractions from control, AsymAD, AD and Parkinson’s disease (PD) brain. Relative levels of specific insoluble RNA-binding proteins across different disease groups correlated with accumulation of Aβ and tau aggregates. RNA-binding proteins, including splicing factors with homology to the basic-acidic dipeptide repeats of U1-70K, preferentially aggregated in AsymAD and AD. In contrast, PD brain aggregates were relatively depleted of many RNA-binding proteins compared to AsymAD and AD groups. Correlation network analyses resolved 29 distinct modules of co-aggregating proteins including modules linked to spliceosome assembly, nuclear speckles and RNA splicing. Modules related to spliceosome assembly and nuclear speckles showed stage-specific enrichment of insoluble RBPs from AsymAD and AD brains, whereas the RNA splicing module was reduced specifically in PD. Collectively, this work identifies classes of RNA-binding proteins that distinctly co-aggregate in detergent-insoluble fractions across the specific neurodegenerative diseases we examined.

scholarly journals Secondary structure of pre-mRNA introns for genes in the 15q11-12 locus. Mapping of functionally significant motives for RNA-binding proteins and nucleosome positioning signals

2018 ◽  
Author(s):  
Viya B. Fedoseyeva ◽  
Irina A. Zharinova ◽  
Alexander A. Alexandrov
Keyword(s):  
Secondary Structure ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Nucleosome Positioning ◽  
Nuclear Speckles ◽  
Dominant Type ◽  
Mrna Variants ◽  
Locus Mapping ◽  
Folded Structures

AbstractIn this study, we identified reproducible substructures in the folded structures of long intron RNAs for recursive spliced variants and annotated pre-mRNA for GABRB3 and GABRA5. We mapped the RNA motives recognized by RNA-binding proteins for the specified locus and characterized the area of preferred localization. A comparison of pre-mRNA variants revealed the dominant type of protein potential effects. We determined the structural specifics of RNA in the dense Alu cluster and clarified the analogy of apical substructure to the A-Xist fragment of transcriptional variant. Mapping of the nucleosome potential reveals alternation of strong and weak signals at the 3’-end portion of GABRB3 and clusters of nucleosome positioning signal in the vicinity of the Alu cluster. Distribution of simple oligonucleotides among reproducible substructures revealed an enrichment in Py-tracts; for some of them, this may be considered as a complementary supplement to the Pu-tract enrichment of ncRNA Malat1 as a component of nuclear speckles. The secondary structure elements of bidirectional transcripts are predisposed for somatic homolog pairing in this locus, as was previously shown experimentally.A model of potential intron RNA influence on splicing has been suggested based on its interaction with Py-tract-binding RNP, serine-arginine SRSF proteins, ncRNA Malat1, as well as the action of Alu cluster.

scholarly journals An interaction network of RNA-binding proteins involved in Drosophila oogenesis

2020 ◽  
Author(s):  
Prashali Bansal ◽  
Johannes Madlung ◽  
Kristina Schaaf ◽  
Boris Macek ◽  
Fulvia Bono
Keyword(s):  
Translational Regulation ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Direct Interaction ◽  
Interaction Network ◽  
Splicing Factors ◽  
Mrna Regulation ◽  
Interaction Partners ◽  
Maternal Transcripts

AbstractDuring Drosophila oogenesis, the localization and translational regulation of maternal transcripts relies on RNA-binding proteins (RBPs). Many of these RBPs localize several mRNAs and may have additional direct interaction partners to regulate their functions. Using immunoprecipitation from whole Drosophila ovaries coupled to mass spectrometry, we examined protein-protein associations of 6 GFP-tagged RBPs expressed at physiological levels. Analysis of the interaction network and further validation in human cells allowed us to identify 26 previously unknown associations, besides recovering several well characterized interactions. We identified interactions between RBPs and several splicing factors, providing links between nuclear and cytoplasmic events of mRNA regulation. Additionally, components of the translational and RNA decay machineries were selectively co-purified with some baits, suggesting a mechanism for how RBPs may regulate maternal transcripts. Given the evolutionary conservation of the studied RBPs, the interaction network presented here provides the foundation for future functional and structural studies of mRNA localization across metazoans.

scholarly journals An Interaction Network of RNA-Binding Proteins Involved in Drosophila Oogenesis

2020 ◽  
Vol 19 (9) ◽  
pp. 1485-1502
Author(s):  
Prashali Bansal ◽  
Johannes Madlung ◽  
Kristina Schaaf ◽  
Boris Macek ◽  
Fulvia Bono
Keyword(s):  
Translational Regulation ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Direct Interaction ◽  
Interaction Network ◽  
Splicing Factors ◽  
Mrna Regulation ◽  
Interaction Partners ◽  
Maternal Transcripts

During Drosophila oogenesis, the localization and translational regulation of maternal transcripts relies on RNA-binding proteins (RBPs). Many of these RBPs localize several mRNAs and may have additional direct interaction partners to regulate their functions. Using immunoprecipitation from whole Drosophila ovaries coupled to mass spectrometry, we examined protein-protein associations of 6 GFP-tagged RBPs expressed at physiological levels. Analysis of the interaction network and further validation in human cells allowed us to identify 26 previously unknown associations, besides recovering several well characterized interactions. We identified interactions between RBPs and several splicing factors, providing links between nuclear and cytoplasmic events of mRNA regulation. Additionally, components of the translational and RNA decay machineries were selectively co-purified with some baits, suggesting a mechanism for how RBPs may regulate maternal transcripts. Given the evolutionary conservation of the studied RBPs, the interaction network presented here provides the foundation for future functional and structural studies of mRNA localization across metazoans.

scholarly journals Pathological tau drives ectopic nuclear speckle scaffold protein SRRM2 accumulation in neuron cytoplasm in Alzheimer’s disease

2021 ◽  
Vol 9 (1) ◽  
Author(s):  
Pamela J. McMillan ◽  
Timothy J. Strovas ◽  
Misa Baum ◽  
Brooke K. Mitchell ◽  
Randall J. Eck ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Transgenic Mice ◽  
Brain Tissue ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Scaffold Protein ◽  
Nuclear Speckles ◽  
Nuclear Speckle ◽  
C Elegans

AbstractSeveral conserved nuclear RNA binding proteins (sut-1, sut-2, and parn-2) control tau aggregation and toxicity in C. elegans, mice, and human cells. MSUT2 protein normally resides in nuclear speckles, membraneless organelles composed of phase-separated RNAs and RNA-binding proteins that mediate critical steps in mRNA processing including mRNA splicing. We used human pathological tissue and transgenic mice to identify Alzheimer’s disease-specific cellular changes related to nuclear speckles. We observed that nuclear speckle constituent scaffold protein SRRM2 is mislocalized and accumulates in cytoplasmic lesions in AD brain tissue. Furthermore, progression of tauopathy in transgenic mice is accompanied by increasing mislocalization of SRRM2 from the neuronal nucleus to the soma. In AD brain tissue, SRRM2 mislocalization associates with increased severity of pathological tau deposition. These findings suggest potential mechanisms by which pathological tau impacts nuclear speckle function in diverse organisms ranging from C. elegans to mice to humans. Future translational studies aimed at restoring nuclear speckle homeostasis may provide novel candidate therapeutic targets for pharmacological intervention.

scholarly journals Identification of Proteins Bound to Dengue Viral RNA In Vivo Reveals New Host Proteins Important for Virus Replication

mBio ◽  
2016 ◽  
Vol 7 (1) ◽  
Author(s):  
Stacia L. Phillips ◽  
Erik J. Soderblom ◽  
Shelton S. Bradrick ◽  
Mariano A. Garcia-Blanco
Keyword(s):  
Mass Spectrometry ◽  
Dengue Virus ◽  
Virus Replication ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Viral Rna ◽  
Host Proteins ◽  
Infected Cells

ABSTRACT Dengue virus is the most prevalent cause of arthropod-borne infection worldwide. Due to the limited coding capacity of the viral genome and the complexity of the viral life cycle, host cell proteins play essential roles throughout the course of viral infection. Host RNA-binding proteins mediate various aspects of virus replication through their physical interactions with viral RNA. Here we describe a technique designed to identify such interactions in the context of infected cells using UV cross-linking followed by antisense-mediated affinity purification and mass spectrometry. Using this approach, we identified interactions, several of them novel, between host proteins and dengue viral RNA in infected Huh7 cells. Most of these interactions were subsequently validated using RNA immunoprecipitation. Using small interfering RNA (siRNA)-mediated gene silencing, we showed that more than half of these host proteins are likely involved in regulating virus replication, demonstrating the utility of this method in identifying biologically relevant interactions that may not be identified using traditional in vitro approaches. IMPORTANCE Dengue virus is the most prevalent cause of arthropod-borne infection worldwide. Viral RNA molecules physically interact with cellular RNA-binding proteins (RBPs) throughout the course of infection; the identification of such interactions will lead to the elucidation of the molecular mechanisms of virus replication. Until now, the identification of host proteins bound to dengue viral RNA has been accomplished using in vitro strategies. Here, we used a method for the specific purification of dengue viral ribonucleoprotein (RNP) complexes from infected cells and subsequently identified the associated proteins by mass spectrometry. We then validated a functional role for the majority of these proteins in mediating efficient virus replication. This approach has broad relevance to virology and RNA biology, as it could theoretically be used to purify any viral RNP complex of interest.

scholarly journals RNA-Binding Proteins at the Host-Pathogen Interface Targeting Viral Regulatory Elements

Viruses ◽  
2021 ◽  
Vol 13 (6) ◽  
pp. 952
Author(s):  
Azman Embarc-Buh ◽  
Rosario Francisco-Velilla ◽  
Encarnacion Martinez-Salas
Keyword(s):  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Rna Viruses ◽  
Regulatory Elements ◽  
Host Cells ◽  
Viral Rnas ◽  
Positive Strand ◽  
Multiplication Cycle ◽  
Positive Strand Rna

Viral RNAs contain the information needed to synthesize their own proteins, to replicate, and to spread to susceptible cells. However, due to their reduced coding capacity RNA viruses rely on host cells to complete their multiplication cycle. This is largely achieved by the concerted action of regulatory structural elements on viral RNAs and a subset of host proteins, whose dedicated function across all stages of the infection steps is critical to complete the viral cycle. Importantly, not only the RNA sequence but also the RNA architecture imposed by the presence of specific structural domains mediates the interaction with host RNA-binding proteins (RBPs), ultimately affecting virus multiplication and spreading. In marked difference with other biological systems, the genome of positive strand RNA viruses is also the mRNA. Here we focus on distinct types of positive strand RNA viruses that differ in the regulatory elements used to promote translation of the viral RNA, as well as in the mechanisms used to evade the series of events connected to antiviral response, including translation shutoff induced in infected cells, assembly of stress granules, and trafficking stress.

scholarly journals RNA-Binding Proteins: Splicing Factors and Disease

Biomolecules ◽  
2015 ◽  
Vol 5 (2) ◽  
pp. 893-909 ◽  
Author(s):  
Alger Fredericks ◽  
Kamil Cygan ◽  
Brian Brown ◽  
William Fairbrother
Keyword(s):  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Splicing Factors
Sign in / Sign up

Export Citation Format

Share Document