scholarly journals HIV-1 RNA genomes initiate host protein packaging in the cytosol independently of Gag capsid proteins

2019 ◽  
Author(s):  
Jordan T. Becker ◽  
Edward L. Evans ◽  
Bayleigh E. Benner ◽  
Stephanie L. Pritzl ◽  
Laura E. Smith ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Host Protein ◽  
Capsid Proteins ◽  
Particle Assembly ◽  
Nonsense Mediated Decay ◽  
Genome Interactions ◽  
Hiv 1

ABSTRACTHIV-1 RNA genomes interact with diverse RNA binding proteins (RBPs) in the cytoplasm including antiviral factor APOBEC3G (A3G) that, in the absence of viral Vif proteins, is packaged into virions. Where and when HIV-1-A3G interactions are initiated for packaging inside the cell is unknown, and the relative contributions of genome vs. Gag capsid proteins to this process remains controversial. Here we visualized A3G responses to HIV-1 infection over an entire replication cycle using long-term (up to 72 h) live single cell imaging. We show that Vif-deficient HIV-1 dramatically shifts A3G and a second RNA surveillance factor, MOV10, from the cytoplasm to virus particle assembly sites with little to no net discernible effects on general markers of cytoplasmic processing bodies (DCP1A), stress granules (TIA-1), or a marker of the nonsense-mediated decay machinery (UPF1). Using a new live cell RNA-protein interaction assay based on RNA tethering (the in-cell RNA-protein interaction protocol, or IC-IP), we provide evidence that A3G- and MOV10- genome interactions are selective, strong, occur in presence or absence of Gag, and are initiated in the cytosol soon if not immediately after genome nuclear export. Finally, although Gag is sufficient to package A3G into virions even in the absence of genomes, single virion imaging indicates that selective A3G-genome interactions promote much more consistent per virion delivery of A3G to assembly sites. Collectively, these studies suggest a paradigm for early, strong, and persistent cytosolic detection of select HIV-1 RNA signatures by A3G, MOV10 and other host RBPs that are enriched in virions.IMPORTANCEHost-pathogen interactions determine the success of viral replication. While extensive work has identified many interactions between HIV-1 and cellular factors, our understanding of where these interactions in cells occur during the course of infection is incomplete. Here, we show that multiple RNA-binding proteins (including the antiviral restriction factor, APOBEC3G, and MOV10 helicase) bind HIV-1 RNA genomes in the cytoplasm and co-traffic with them into progeny virions. Furthermore, we show that these interactions with HIV-1 RNA occur in the absence of Gag and are sufficiently strong to recruit these to otherwise non-native subcellular locales. Together, these data begin to illuminate the intracellular trafficking pathways shared by host RNA binding proteins and the viral RNAs they preferentially bind.

scholarly journals RNA-binding proteins of the NXF (nuclear export factor) family and their connection with the cytoskeleton

Cytoskeleton ◽  
2017 ◽  
Vol 74 (4) ◽  
pp. 161-169 ◽  
Author(s):  
L. A. Mamon ◽  
V. R. Ginanova ◽  
S. F. Kliver ◽  
A. O. Yakimova ◽  
A. A. Atsapkina ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins

scholarly journals Matrin3: Disorder and ALS Pathogenesis

2022 ◽  
Vol 8 ◽  
Author(s):  
Ahmed Salem ◽  
Carter J. Wilson ◽  
Benjamin S. Rutledge ◽  
Allison Dilliott ◽  
Sali Farhan ◽  
...  
Keyword(s):  
Nuclear Export ◽  
Binding Proteins ◽  
Rna Binding ◽  
Nuclear Dna ◽  
Neurodegenerative Disorder ◽  
Rna Binding Proteins ◽  
Binding Protein ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Disordered Regions

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder characterized by the degeneration of both upper and lower motor neurons in the brain and spinal cord. ALS is associated with protein misfolding and inclusion formation involving RNA-binding proteins, including TAR DNA-binding protein (TDP-43) and fused in sarcoma (FUS). The 125-kDa Matrin3 is a highly conserved nuclear DNA/RNA-binding protein that is implicated in many cellular processes, including binding and stabilizing mRNA, regulating mRNA nuclear export, modulating alternative splicing, and managing chromosomal distribution. Mutations in MATR3, the gene encoding Matrin3, have been identified as causal in familial ALS (fALS). Matrin3 lacks a prion-like domain that characterizes many other ALS-associated RNA-binding proteins, including TDP-43 and FUS, however, our bioinformatics analyses and preliminary studies document that Matrin3 contains long intrinsically disordered regions that may facilitate promiscuous interactions with many proteins and may contribute to its misfolding. In addition, these disordered regions in Matrin3 undergo numerous post-translational modifications, including phosphorylation, ubiquitination and acetylation that modulate the function and misfolding of the protein. Here we discuss the disordered nature of Matrin3 and review the factors that may promote its misfolding and aggregation, two elements that might explain its role in ALS pathogenesis.

HIV-1 TAR RNA-binding proteins control TAT activation of translation in Xenopus oocytes.

1993 ◽  
Vol 7 (1) ◽  
pp. 214-222 ◽  
Author(s):  
M Braddock ◽  
R Powell ◽  
A D Blanchard ◽  
A J Kingsman ◽  
S M Kingsman
Keyword(s):  
Xenopus Oocytes ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Tar Rna ◽  
Hiv 1

scholarly journals HIV-1 remodels the nuclear pore complex

2011 ◽  
Vol 193 (4) ◽  
pp. 619-631 ◽  
Author(s):  
Anne Monette ◽  
Nelly Panté ◽  
Andrew J. Mouland
Keyword(s):  
Nuclear Export ◽  
Cell Biology ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Immunogold Electron Microscopy ◽  
Nuclear Pore ◽  
Viral Control ◽  
Host Cell Proteins ◽  
Associated Proteins ◽  
Hiv 1

Human immunodeficiency virus type 1 (HIV-1) commandeers host cell proteins and machineries for its replication. Our earlier work showed that HIV-1 induced the cytoplasmic retention of nucleocytoplasmic shuttling and ribonucleic acid (RNA)–binding proteins. This retention is dependent on nuclear export of the viral genomic RNA and on changes in the localization and expression level of the nucleoporin (Nup) p62 (Nup62). To further characterize the extent of perturbation induced by HIV-1, we performed proteomics analyses of nuclear envelopes (NEs) isolated from infected T cells. Infection induced extensive changes in the composition of the NE and its associated proteins, including a remarkable decrease in the abundance of Nups. Immunogold electron microscopy revealed the translocation of Nups into the cytoplasm. Nup62 was identified as a component of purified virus, and small interfering RNA depletion studies revealed an important role for this Nup in virus gene expression and infectivity. This detailed analysis highlights the profound effects on NE composition induced by HIV-1 infection, providing further evidence of the magnitude of viral control over the cell biology of its host.

scholarly journals Trypanosoma brucei RNA Binding Proteins p34 and p37 Mediate NOPP44/46 Cellular Localization via the Exportin 1 Nuclear Export Pathway

2007 ◽  
Vol 6 (12) ◽  
pp. 2206-2213 ◽  
Author(s):  
Kristina Hellman ◽  
Kimberly Prohaska ◽  
Noreen Williams
Keyword(s):  
Amino Acid ◽  
Trypanosoma Brucei ◽  
Nuclear Export ◽  
Cellular Localization ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Nuclear Export Signal ◽  
Amino Acid Sequences ◽  
Protein Levels

ABSTRACT We have previously identified and characterized two novel nuclear RNA binding proteins, p34 and p37, which have been shown to interact with a family of nucleolar phosphoproteins, NOPP44/46, in Trypanosoma brucei. These proteins are nearly identical, the major difference being an 18-amino-acid insert in the N terminus of p37. In order to characterize the interaction between p34 and p37 and NOPP44/46, we have utilized an RNA interference (RNAi) cell line that specifically targets p34 and p37. Within these RNAi cells, we detected a disruption of a higher-molecular-weight complex containing NOPP44/46, as well as a dramatic increase in nuclear NOPP44/46 protein levels. We demonstrated that no change occurred in NOPP44/46 mRNA steady-state levels or stability, nor was there a change in cellular protein levels. These results led us to investigate whether p34 and p37 regulate NOPP44/46 cellular localization. Examination of the p34 and p37 amino acid sequences revealed a leucine-rich nuclear export signal, which interacts with the nuclear export factor exportin 1. Immune capture experiments demonstrated that p34, p37, and NOPP44/46 associate with exportin 1. When these experiments were performed with p34/p37 RNAi cells, NOPP44/46 no longer associated with exportin 1. Sequential immune capture experiments demonstrated that p34, p37, NOPP44/46, and exportin 1 exist in a common complex. Inhibiting exportin 1-mediated nuclear export led to an increase in nuclear NOPP44/46 proteins, indicating that they are exported from the nucleus via this pathway. Together, our results demonstrate that p34 and p37 regulate NOPP44/46 cellular localization by facilitating their association with exportin 1.

scholarly journals Abstract P-22: Enhanced Crosslinking and Immunoprecipitation (Eclip) Data Reveal Interactions of RNA Binding Proteins with the Human Ribosome

2021 ◽  
Vol 11 (Suppl_1) ◽  
pp. S21-S21
Author(s):  
Andrey Buyan ◽  
Ivan Kulakovskiy ◽  
Sergey Dmitriev
Keyword(s):  
Translational Control ◽  
Nuclear Export ◽  
Ribosomal Proteins ◽  
Ribosome Biogenesis ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Repetitive Sequences ◽  
Structural Data

Background: The ribosome is a protein-synthesizing molecular machine composed of four ribosomal RNAs (rRNAs) and dozens of ribosomal proteins. In mammals, the ribosome has a complicated structure with an additional outer layer of rRNA, including large tentacle-like extensions. A number of RNA binding proteins (RBPs) interact with this layer to assist ribosome biogenesis, nuclear export and decay, or to modulate translation. Plenty of methods have been developed in the last decade in order to study such protein-RNA interactions, including RNA pulldown and crosslinking-immunoprecipitation (CLIP) assays. Methods: In the current study, using publicly available data of the enhanced CLIP (eCLIP) experiments for 223 proteins studied in the ENCODE project, we found a number of RBPs that bind rRNAs in human cells. To locate their binding sites in rRNAs, we used a newly developed computational protocol for mapping and evaluation of the eCLIP data with the respect to the repetitive sequences. Results: For two proteins with known ribosomal localization, uS3/RPS3 and uS17/RPS11, the identified sites were in good agreement with structural data, thus validating our approach. Then, we identified rRNA contacts of overall 22 RBPs involved in rRNA processing and ribosome maturation (DDX21, DDX51, DDX52, NIP7, SBDS, UTP18, UTP3, WDR3, and WDR43), translational control during stress (SERBP1, G3BP1, SND1), IRES activity (PCBP1/hnRNPE1), and other translation-related functions. In many cases, the identified proteins interact with the rRNA expansion segments (ES) of the human ribosome pointing to their important role in protein synthesis. Conclusion: Our study identifies a number of RBPs as interacting partners of the human ribosome and sheds light on the role of rRNA expansion segments in translation.

scholarly journals Systematic discovery and functional interrogation of SARS-CoV-2 viral RNA-host protein interactions during infection

2020 ◽  
Author(s):  
Ryan A. Flynn ◽  
Julia A. Belk ◽  
Yanyan Qi ◽  
Yuki Yasumoto ◽  
Cameron O. Schmitz ◽  
...  
Keyword(s):  
Protein Interaction ◽  
Protein Interactions ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Viral Rna ◽  
Host Protein ◽  
List Type ◽  
Genome Wide ◽  
A Genome

AbstractSevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the cause of a pandemic with growing global mortality. There is an urgent need to understand the molecular pathways required for host infection and anti-viral immunity. Using comprehensive identification of RNA-binding proteins by mass spectrometry (ChIRP-MS), we identified 309 host proteins that bind the SARS-CoV-2 RNA during active infection. Integration of this data with viral ChIRP-MS data from three other positive-sense RNA viruses defined pan-viral and SARS-CoV-2-specific host interactions. Functional interrogation of these factors with a genome-wide CRISPR screen revealed that the vast majority of viral RNA-binding proteins protect the host from virus-induced cell death, and we identified known and novel anti-viral proteins that regulate SARS-CoV-2 pathogenicity. Finally, our RNA-centric approach demonstrated a physical connection between SARS-CoV-2 RNA and host mitochondria, which we validated with functional and electron microscopy data, providing new insights into a more general virus-specific protein logic for mitochondrial interactions. Altogether, these data provide a comprehensive catalogue of SARS-CoV-2 RNA-host protein interactions, which may inform future studies to understand the mechanisms of viral pathogenesis, as well as nominate host pathways that could be targeted for therapeutic benefit.Highlights· ChIRP-MS of SARS-CoV-2 RNA identifies a comprehensive viral RNA-host protein interaction network during infection across two species· Comparison to RNA-protein interaction networks with Zika virus, dengue virus, and rhinovirus identify SARS-CoV-2-specific and pan-viral RNA protein complexes and highlights distinct intracellular trafficking pathways· Intersection of ChIRP-MS and genome-wide CRISPR screens identify novel SARS-CoV-2-binding proteins with pro- and anti-viral function· Viral RNA-RNA and RNA-protein interactions reveal specific SARS-CoV-2-mediated mitochondrial dysfunction during infection

scholarly journals The RNA-binding protein PTBP1 promotes ATPase-dependent dissociation of the RNA helicase UPF1 to protect transcripts from nonsense-mediated mRNA decay

2020 ◽  
Vol 295 (33) ◽  
pp. 11613-11625 ◽  
Author(s):  
Sarah E. Fritz ◽  
Soumya Ranganathan ◽  
Clara D. Wang ◽  
J. Robert Hogg
Keyword(s):  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Binding Protein ◽  
Target Selection ◽  
Rna Helicase ◽  
Nonsense Mediated Decay ◽  
Polypyrimidine Tract Binding Protein ◽  
Regulatory Loop

The sequence-specific RNA-binding proteins PTBP1 (polypyrimidine tract–binding protein 1) and HNRNP L (heterogeneous nuclear ribonucleoprotein L) protect mRNAs from nonsense-mediated decay (NMD) by preventing the UPF1 RNA helicase from associating with potential decay targets. Here, by analyzing in vitro helicase activity, dissociation of UPF1 from purified mRNPs, and transcriptome-wide UPF1 RNA binding, we present the mechanistic basis for inhibition of NMD by PTBP1. Unlike mechanisms of RNA stabilization that depend on direct competition for binding sites among protective RNA-binding proteins and decay factors, PTBP1 promotes displacement of UPF1 already bound to potential substrates. Our results show that PTBP1 directly exploits the tendency of UPF1 to release RNA upon ATP binding and hydrolysis. We further find that UPF1 sensitivity to PTBP1 is coordinated by a regulatory loop in domain 1B of UPF1. We propose that the UPF1 regulatory loop and protective proteins control kinetic proofreading of potential NMD substrates, presenting a new model for RNA helicase regulation and target selection in the NMD pathway.

scholarly journals Exportin-5, a novel karyopherin, mediates nuclear export of double-stranded RNA binding proteins

2002 ◽  
Vol 156 (1) ◽  
pp. 53-64 ◽  
Author(s):  
Amy M. Brownawell ◽  
Ian G. Macara
Keyword(s):  
Nuclear Export ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Dependent Manner ◽  
Intact Cells ◽  
Double Stranded Rna ◽  
Permeabilized Cells ◽  
Dsrna Binding ◽  
Rna Export

We have identified a novel human karyopherin (Kap)β family member that is related to human Crm1 and the Saccharomyces cerevisiae protein, Msn5p/Kap142p. Like other known transport receptors, this Kap binds specifically to RanGTP, interacts with nucleoporins, and shuttles between the nuclear and cytoplasmic compartments. We report that interleukin enhancer binding factor (ILF)3, a double-stranded RNA binding protein, associates with this Kap in a RanGTP-dependent manner and that its double-stranded RNA binding domain (dsRBD) is the limiting sequence required for this interaction. Importantly, the Kap interacts with dsRBDs found in several other proteins and binding is blocked by double-stranded RNA. We find that the dsRBD of ILF3 functions as a novel nuclear export sequence (NES) in intact cells, and its ability to serve as an NES is dependent on the expression of the Kap. In digitonin-permeabilized cells, the Kap but not Crm1 stimulated nuclear export of ILF3. Based on the ability of this Kap to mediate the export of dsRNA binding proteins, we named the protein exportin-5. We propose that exportin-5 is not an RNA export factor but instead participates in the regulated translocation of dsRBD proteins to the cytoplasm where they interact with target mRNAs.

scholarly journals HIV-1 and two avian retroviral 5′ untranslated regions bind orthologous human and chicken RNA binding proteins

Virology ◽  
2015 ◽  
Vol 486 ◽  
pp. 307-320 ◽  
Author(s):  
Matthew Stake ◽  
Deepali Singh ◽  
Gatikrushna Singh ◽  
J. Marcela Hernandez ◽  
Rebecca Kaddis Maldonado ◽  
...  
Keyword(s):  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Untranslated Regions ◽  
Hiv 1
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