scholarly journals RBBP6 activates the pre-mRNA 3′-end processing machinery in humans

2021 ◽  
Author(s):  
Vytaute Boreikaite ◽  
Thomas Elliot ◽  
Jason Chin ◽  
Lori A Passmore
Keyword(s):  
Rna Processing ◽  
Mrna Localization ◽  
Dependent Manner ◽  
Endonucleolytic Cleavage ◽  
Mature Transcript ◽  
Cleavage And Polyadenylation ◽  
Specificity Factor ◽  
Additional Protein ◽  
Stimulatory Factor

3′-end processing of most human mRNAs is carried out by the cleavage and polyadenylation specificity factor (CPSF; CPF in yeast). Endonucleolytic cleavage of the nascent pre-mRNA defines the 3′-end of the mature transcript, which is important for mRNA localization, translation and stability. Cleavage must therefore be tightly regulated. Here, we reconstitute specific and efficient 3′-endonuclease activity of human CPSF with purified proteins. This requires the seven-subunit CPSF as well as three additional protein factors: cleavage stimulatory factor (CStF), cleavage factor IIm (CFIIm) and, importantly, the multi-domain protein RBBP6. Unlike its yeast homologue Mpe1, which is a stable subunit of CPF, RBBP6 does not copurify with CPSF and is recruited in an RNA-dependent manner. Sequence and mutational analyses suggest that RBBP6 interacts with the WDR33 and CPSF73 subunits of CPSF. Thus, it is likely that the role of RBBP6 is conserved from yeast to human. Overall, our data are consistent with CPSF endonuclease activation and site-specific pre-mRNA cleavage being highly controlled to maintain fidelity in RNA processing.

scholarly journals Cleavage and Polyadenylation Specificity Factor 6 Is Required for Efficient HIV-1 Latency Reversal

mBio ◽  
2021 ◽  
Author(s):  
Yue Zheng ◽  
Heidi L. Schubert ◽  
Parmit K. Singh ◽  
Laura J. Martins ◽  
Alan N. Engelman ◽  
...  
Keyword(s):  
Life Cycle ◽  
Protein Phosphatase ◽  
Pol Ii ◽  
Phosphatase 2A ◽  
Cleavage And Polyadenylation ◽  
Specificity Factor ◽  
Critical Residues ◽  
The Stability ◽  
Hiv 1

CPSF6 is a cellular factor that regulates cleavage and polyadenylation of mRNAs and participates in HIV-1 infection by facilitating targeting of preintegration complexes to the chromatin. Our observations reveal a second role of CPSF6 in the HIV-1 life cycle that involves regulation of viral transcription through controlling the stability of protein phosphatase 2A, which in turn regulates the phosphorylation/dephosphorylation status of critical residues in CDK9 and Pol II.

scholarly journals Role of poly(A) polymerase in the cleavage and polyadenylation of mRNA precursor.

1989 ◽  
Vol 9 (4) ◽  
pp. 1435-1444 ◽  
Author(s):  
M P Terns ◽  
S T Jacob
Keyword(s):  
Cleavage Site ◽  
Cleavage Reaction ◽  
Endonucleolytic Cleavage ◽  
Nuclear Extracts ◽  
Adenylic Acid ◽  
Specific Complex ◽  
Functional Complex ◽  
Cleavage And Polyadenylation ◽  
Mrna Precursor

To determine the role of poly(A) polymerase in 3'-end processing of mRNA, the effect of purified poly(A) polymerase antibodies on endonucleolytic cleavage and polyadenylation was studied in HeLa nuclear extracts, using adenovirus L3 pre-mRNA as the substrate. Both Mg2+- and Mn2+-dependent reactions catalyzing addition of 200 to 250 and 400 to 800 adenylic acid residues, respectively, were inhibited by the antibodies, which suggested that the two reactions were catalyzed by the same enzyme. Anti-poly(A) polymerase antibodies also inhibited the cleavage reaction when the reaction was coupled or chemically uncoupled with polyadenylation. These antibodies also prevented formation of specific complexes between the RNA substrate and components of nuclear extracts during cleavage or polyadenylation, with the concurrent appearance of another, antibody-specific complex. These studies demonstrate that (i) previously characterized poly(A) polymerase is the enzyme responsible for addition of the poly(A) tract at the correct cleavage site and probably for the elongation of poly(A) chains and (ii) the coupling of these two 3'-end processing reactions appears to result from the potential requirement of poly(A) polymerase for the cleavage reaction. The results suggest that the specific endonuclease is associated with poly(A) polymerase in a functional complex.

scholarly journals Cytoplasmic CPSF6 Regulates HIV-1 Capsid Trafficking and Infection in a Cyclophilin A-Dependent Manner

mBio ◽  
2021 ◽  
Vol 12 (2) ◽  
Author(s):  
Zhou Zhong ◽  
Jiying Ning ◽  
Emerson A. Boggs ◽  
Sooin Jang ◽  
Callen Wallace ◽  
...  
Keyword(s):  
T Cells ◽  
Cell Nucleus ◽  
Viral Genome ◽  
Cyclophilin A ◽  
Higher Order ◽  
Human Protein ◽  
Dependent Manner ◽  
Cleavage And Polyadenylation ◽  
Specificity Factor ◽  
Hiv 1

ABSTRACT Human immunodeficiency virus type 1 (HIV-1) capsid binds host proteins during infection, including cleavage and polyadenylation specificity factor 6 (CPSF6) and cyclophilin A (CypA). We observe that HIV-1 infection induces higher-order CPSF6 formation, and capsid-CPSF6 complexes cotraffic on microtubules. CPSF6-capsid complex trafficking is impacted by capsid alterations that reduce CPSF6 binding or by excess cytoplasmic CPSF6 expression, both of which are associated with decreased HIV-1 infection. Higher-order CPSF6 complexes bind and disrupt HIV-1 capsid assemblies in vitro. Disruption of HIV-1 capsid binding to CypA leads to increased CPSF6 binding and altered capsid trafficking, resulting in reduced infectivity. Our data reveal an interplay between CPSF6 and CypA that is important for cytoplasmic capsid trafficking and HIV-1 infection. We propose that CypA prevents HIV-1 capsid from prematurely engaging cytoplasmic CPSF6 and that differences in CypA cellular localization and innate immunity may explain variations in HIV-1 capsid trafficking and uncoating in CD4+ T cells and macrophages. IMPORTANCE HIV is the causative agent of AIDS, which has no cure. The protein shell that encases the viral genome, the capsid, is critical for HIV replication in cells at multiple steps. HIV capsid has been shown to interact with multiple cell proteins during movement to the cell nucleus in a poorly understood process that may differ during infection of different cell types. In this study, we show that premature or too much binding of one human protein, cleavage and polyadenylation specificity factor 6 (CPSF6), disrupts the ability of the capsid to deliver the viral genome to the cell nucleus. Another human protein, cyclophilin A (CypA), can shield HIV capsid from premature binding to CPSF6, which can differ in CD4+ T cells and macrophages. Better understanding of how HIV infects cells will allow better drugs to prevent or inhibit infection and pathogenesis.

scholarly journals Cytoplasmic CPSF6 regulates HIV-1 capsid trafficking and infection in a cyclophilin A-dependent manner

2020 ◽  
Author(s):  
Zhou Zhong ◽  
Jiying Ning ◽  
Emerson A. Boggs ◽  
Sooin Jang ◽  
Callen Wallace ◽  
...  
Keyword(s):  
Cellular Localization ◽  
Cyclophilin A ◽  
Higher Order ◽  
Dependent Manner ◽  
Host Proteins ◽  
Immunodeficiency Virus ◽  
Cleavage And Polyadenylation ◽  
Specificity Factor ◽  
Hiv 1

SummaryHuman immunodeficiency virus type 1 (HIV-1) capsid binds host proteins during infection, including cleavage and polyadenylation specificity factor 6 (CPSF6) and cyclophilin A (CypA). We observe that HIV-1 infection induces higher-order CPSF6 formation and capsid-CPSF6 complexes co-traffic on microtubules. CPSF6-capsid complex trafficking is impacted by capsid alterations that reduce CPSF6 binding or by excess cytoplasmic CPSF6 expression, both of which are associated with decreased HIV-1 infection. Higher-order CPSF6 complexes bind and disrupt HIV-1 capsid assemblies in vitro. Disruption of HIV-1 capsid binding to CypA leads to increased CPSF6 binding and altered capsid trafficking, resulting in reduced infectivity. Our data reveal an interplay between CPSF6 and CypA that is important for cytoplasmic capsid trafficking and HIV-1 infection. We propose that CypA prevents HIV-1 capsid from prematurely engaging cytoplasmic CPSF6 and that differences in CypA cellular localization and innate immunity may explain cell-specific variations in HIV-1 capsid trafficking and uncoating.Graphical Abstract

Abstract 17448: Comprehensive Characterization of Alternative Polyadenylation in Human Right Ventricle Failure

Circulation ◽  
2020 ◽  
Vol 142 (Suppl_3) ◽  
Author(s):  
Rajarajan Amirthalingam Thandavarayan ◽  
Scott D Collum ◽  
Keith Youker ◽  
Rajaseakaran Mahalingam ◽  
Leng Han ◽  
...  
Keyword(s):  
Right Ventricle ◽  
Rna Processing ◽  
Alternative Polyadenylation ◽  
Specific Gene ◽  
Gene Expressions ◽  
Human Right ◽  
Additional Layer ◽  
Cleavage And Polyadenylation ◽  
Specificity Factor ◽  
The Right

Background: Alternative cleavage and polyadenylation (APA) an emerging post-transcriptional mechanism for gene regulation that generates distinct isoforms of mRNA with different 3’UTR lengths. APA plays an important role in different biological processes and dysregulation of APA leading to many human diseases. However, the functional consequences of APA events in the right ventricle (RV) failure in humans remain unexplored. Objective: To identify whether the 3′UTR length is modulated by APA in the RV failure in humans compared to healthy RV. Methods and Results: We used Poly A tail RNA sequencing and a novel algorithm called DaPars to measure the global patterns of APA in healthy and failing human RV specimens. We revealed 3'UTR shortening and lengthening of many genes in failing RV compared to healthy RV specimens. The lengthened 3’UTR genes were enriched for functional groups such as RNA processing and splicing. Whereas the shortened 3’UTR genes were enriched for nucleic acid metabolic process and chromatin organization. In addition, differential APA events in RV failure regulates many pathways important for the progression of the RV failure. Finally, the regulator proteins of APA including cleavage and polyadenylation specificity factor (CPSF) 6 and 7, cleavage factor Im (CFIm) 25 and 59 have been regulated in RV failure compared to healthy RV specimens. Conclusions: Our results highlight important roles of APA in the progression of RV failure in humans, including RNA processing, splicing, and specific gene expression. Demonstrating that APA mediated 3’UTR length regulation provides the additional layer of gene expressions in RV failure.

scholarly journals Role of cleavage and polyadenylation specificity factor 100: anchoring poly(A) sites and modulating transcription termination

2017 ◽  
Vol 91 (5) ◽  
pp. 829-839 ◽  
Author(s):  
Juncheng Lin ◽  
Ruqiang Xu ◽  
Xiaohui Wu ◽  
Yingjia Shen ◽  
Qingshun Q. Li
Keyword(s):  
Transcription Termination ◽  
Cleavage And Polyadenylation ◽  
Specificity Factor

Role of poly(A) polymerase in the cleavage and polyadenylation of mRNA precursor

1989 ◽  
Vol 9 (4) ◽  
pp. 1435-1444
Author(s):  
M P Terns ◽  
S T Jacob
Keyword(s):  
Cleavage Site ◽  
Cleavage Reaction ◽  
Endonucleolytic Cleavage ◽  
Nuclear Extracts ◽  
Adenylic Acid ◽  
Specific Complex ◽  
Functional Complex ◽  
Cleavage And Polyadenylation ◽  
Mrna Precursor

To determine the role of poly(A) polymerase in 3'-end processing of mRNA, the effect of purified poly(A) polymerase antibodies on endonucleolytic cleavage and polyadenylation was studied in HeLa nuclear extracts, using adenovirus L3 pre-mRNA as the substrate. Both Mg2+- and Mn2+-dependent reactions catalyzing addition of 200 to 250 and 400 to 800 adenylic acid residues, respectively, were inhibited by the antibodies, which suggested that the two reactions were catalyzed by the same enzyme. Anti-poly(A) polymerase antibodies also inhibited the cleavage reaction when the reaction was coupled or chemically uncoupled with polyadenylation. These antibodies also prevented formation of specific complexes between the RNA substrate and components of nuclear extracts during cleavage or polyadenylation, with the concurrent appearance of another, antibody-specific complex. These studies demonstrate that (i) previously characterized poly(A) polymerase is the enzyme responsible for addition of the poly(A) tract at the correct cleavage site and probably for the elongation of poly(A) chains and (ii) the coupling of these two 3'-end processing reactions appears to result from the potential requirement of poly(A) polymerase for the cleavage reaction. The results suggest that the specific endonuclease is associated with poly(A) polymerase in a functional complex.

Amelioration of Carbon Tetrachloride-Induced Liver Injury by Citrus Leaf Extract

2019 ◽  
Vol 17 (4) ◽  
pp. 426-431
Author(s):  
Jin Xuezhu ◽  
Li Jitong ◽  
Nie Leigang ◽  
Xue Junlai
Keyword(s):  
Carbon Tetrachloride ◽  
Leaf Extract ◽  
Hepatic Injury ◽  
The Body ◽  
Dependent Manner ◽  
Weight Changes ◽  
Citrus Leaf ◽  
Dose Dependent ◽  
And Oxidative Stress

The main purpose of this study is to investigate the role of citrus leaf extract in carbon tetrachloride-induced hepatic injury and its potential molecular mechanism. Carbon tetrachloride was used to construct hepatic injury animal model. To this end, rats were randomly divided into 4 groups: control, carbon tetrachloride-treated, and two carbon tetrachloride + citrus leaf extract-treated groups. The results show that citrus leaf extract treatment significantly reversed the effects of carbon tetrachloride on the body weight changes and liver index. Besides, treatment with citrus leaf extract also reduced the levels of serum liver enzymes and oxidative stress in a dose-dependent manner. H&E staining and western blotting suggested that citrus leaf extract could repair liver histological damage by regulating AMPK and Nrf-2.

The Role of Human T-Lymphotropic Virus Type 1 (HTLV-1)-Infected Dendritic Cells in the Development of HTLV-1-Associated Myelopathy/Tropical Spastic Paraparesis

1999 ◽  
Vol 73 (6) ◽  
pp. 4575-4581 ◽  
Author(s):  
Masahiko Makino ◽  
Satoshi Shimokubo ◽  
Shin-Ichi Wakamatsu ◽  
Shuji Izumo ◽  
Masanori Baba
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
Virus Type ◽  
Spastic Paraparesis ◽  
Tropical Spastic Paraparesis ◽  
Normal Donor ◽  
Dependent Manner ◽  
T Lymphotropic Virus

ABSTRACT The development of human T-lymphotropic virus type 1 (HTLV-1)-associated myelopathy/tropical spastic paraparesis (HAM/TSP) is closely associated with the activation of T cells which are HTLV-1 specific but may cross-react with neural antigens (Ags). Immature dendritic cells (DCs), differentiated from normal donor monocytes by using recombinant granulocyte-macrophage colony-stimulating factor and recombinant interleukin-4, were pulsed with HTLV-1 in vitro. The pulsed DCs contained HTLV-1 proviral DNA and expressed HTLV-1 Gag Ag on their surface 6 days after infection. The DCs matured by lipopolysaccharides stimulated autologous CD4+ T cells and CD8+ T cells in a viral dose-dependent manner. However, the proliferation level of CD4+ T cells was five- to sixfold higher than that of CD8+ T cells. In contrast to virus-infected DCs, DCs pulsed with heat-inactivated virions activated only CD4+ T cells. To clarify the role of DCs in HAM/TSP development, monocytes from patients were cultured for 4 days in the presence of the cytokines. The expression of CD86 Ag on DCs was higher and that of CD1a Ag was more down-regulated than in DCs generated from normal monocytes. DCs from two of five patients expressed HTLV-1 Gag Ag. Furthermore, both CD4+ and CD8+ T cells from the patients were greatly stimulated by contact with autologous DCs pulsed with inactivated viral Ag as well as HTLV-1-infected DCs. These results suggest that DCs are susceptible to HTLV-1 infection and that their cognate interaction with T cells may contribute to the development of HAM/TSP.

scholarly journals Role of MDA5 in regulating CXCL10 expression induced by TLR3 signaling in human rheumatoid fibroblast-like synoviocytes

2021 ◽  
Author(s):  
Tatsuro Saruga ◽  
Tadaatsu Imaizumi ◽  
Shogo Kawaguchi ◽  
Kazuhiko Seya ◽  
Tomoh Matsumiya ◽  
...  
Keyword(s):  
Inflammatory Responses ◽  
Neutralizing Antibody ◽  
Poly I:C ◽  
Enzyme Linked Immunosorbent Assay ◽  
Dependent Manner ◽  
Inflammatory Reactions ◽  
Polyinosinic:Polycytidylic Acid ◽  
Tlr3 Signaling ◽  
Fibroblast Like Synoviocytes

AbstractC-X-C motif chemokine 10 (CXCL10) is an inflammatory chemokine and a key molecule in the pathogenesis of rheumatoid arthritis (RA). Melanoma differentiation-associated gene 5 (MDA5) is an RNA helicase that plays a role in innate immune and inflammatory reactions. The details of the regulatory mechanisms of CXCL10 production and the precise role of MDA5 in RA synovitis have not been fully elucidated. The aim of this study was to examine the role of MDA5 in regulating CXCL10 expression in cultured human rheumatoid fibroblast-like synoviocytes (RFLS). RFLS was stimulated with Toll-like receptor 3 (TLR3) ligand polyinosinic:polycytidylic acid (poly I:C), a synthetic double-stranded RNA mimetic. Expression of interferon beta (IFN-β), MDA5, and CXCL10 was measured by real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR), western blotting, and enzyme-linked immunosorbent assay. A neutralizing antibody of IFN-β and siRNA-mediated MDA5 knockdown were used to determine the role of these molecules in regulating CXCL10 expression downstream of TLR3 signaling in RFLS. Poly I:C induced IFN-β, MDA5, and CXCL10 expression in a concentration- and time-dependent manner. IFN-β neutralizing antibody suppressed the expression of MDA5 and CXCL10, and knockdown of MDA5 decreased a part of CXCL10 expression (p < 0.001). The TLR3/IFN-β/CXCL10 axis may play a crucial role in the inflammatory responses in RA synovium, and MDA5 may be partially involved in this axis.

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