scholarly journals NF-κB activation is a turn on for vaccinia virus phosphoprotein A49 to turn off NF-κB activation

2019 ◽  
Vol 116 (12) ◽  
pp. 5699-5704 ◽  
Author(s):  
Sarah Neidel ◽  
Hongwei Ren ◽  
Alice A. Torres ◽  
Geoffrey L. Smith
Keyword(s):  
Vaccinia Virus ◽  
Ubiquitin Ligase ◽  
Molecular Mimicry ◽  
E3 Ubiquitin Ligase ◽  
Virus Protein ◽  
Biological Regulation ◽  
Turn On ◽  
N Terminus ◽  
Iκbα Phosphorylation ◽  
Necessary And Sufficient

Vaccinia virus protein A49 inhibits NF-κB activation by molecular mimicry and has a motif near the N terminus that is conserved in IκBα, β-catenin, HIV Vpu, and some other proteins. This motif contains two serines, and for IκBα and β-catenin, phosphorylation of these serines enables recognition by the E3 ubiquitin ligase β-TrCP. Binding of IκBα and β-catenin by β-TrCP causes their ubiquitylation and thereafter proteasome-mediated degradation. In contrast, HIV Vpu and VACV A49 are not degraded. This paper shows that A49 is phosphorylated at serine 7 but not serine 12 and that this is necessary and sufficient for binding β-TrCP and antagonism of NF-κB. Phosphorylation of A49 S7 occurs when NF-κB signaling is activated by addition of IL-1β or overexpression of TRAF6 or IKKβ, the kinase needed for IκBα phosphorylation. Thus, A49 shows beautiful biological regulation, for it becomes an NF-κB antagonist upon activation of NF-κB signaling. The virulence of viruses expressing mutant A49 proteins or lacking A49 (vΔA49) was tested. vΔA49 was attenuated compared with WT, but viruses expressing A49 that cannot bind β-TrCP or bind β-TrCP constitutively had intermediate virulence. So A49 promotes virulence by inhibiting NF-κB activation and by another mechanism independent of S7 phosphorylation and NF-κB antagonism. Last, a virus lacking A49 was more immunogenic than the WT virus.

Molecular mimicry of NF-κB by vaccinia virus protein enables selective inhibition of antiviral responses

2021 ◽  
Author(s):  
Jonas D. Albarnaz ◽  
Hongwei Ren ◽  
Alice A. Torres ◽  
Evgeniya V. Shmeleva ◽  
Carlos A. Melo ◽  
...  
Keyword(s):  
Vaccinia Virus ◽  
Molecular Mimicry ◽  
Selective Inhibition ◽  
Virus Protein ◽  
Antiviral Responses

scholarly journals How cyclin A destruction escapes the spindle assembly checkpoint

2010 ◽  
Vol 190 (4) ◽  
pp. 501-509 ◽  
Author(s):  
Barbara Di Fiore ◽  
Jonathon Pines
Keyword(s):  
Ubiquitin Ligase ◽  
Spindle Assembly Checkpoint ◽  
Spindle Assembly ◽  
Cyclin A ◽  
Cyclin Dependent Kinase ◽  
Anaphase Promoting Complex ◽  
N Terminus ◽  
Specific Proteins ◽  
Necessary And Sufficient

The anaphase-promoting complex/cyclosome (APC/C) is the ubiquitin ligase essential to mitosis, which ensures that specific proteins are degraded at specific times to control the order of mitotic events. The APC/C coactivator, Cdc20, is targeted by the spindle assembly checkpoint (SAC) to restrict APC/C activity until metaphase, yet early substrates, such as cyclin A, are degraded in the presence of the active checkpoint. Cdc20 and the cyclin-dependent kinase cofactor, Cks, are required for cyclin A destruction, but how they enable checkpoint-resistant destruction has not been elucidated. In this study, we answer this problem: we show that the N terminus of cyclin A binds directly to Cdc20 and with sufficient affinity that it can outcompete the SAC proteins. Subsequently, the Cks protein is necessary and sufficient to promote cyclin A degradation in the presence of an active checkpoint by binding cyclin A–Cdc20 to the APC/C.

scholarly journals NatB modulates Rb mutant cell death and tumor growth by regulating EGFR/MAPK signaling through the N-end rule pathways

2019 ◽  
Author(s):  
Zhentao Sheng ◽  
Wei Du
Keyword(s):  
Tumor Growth ◽  
Ubiquitin Ligase ◽  
Mapk Pathway ◽  
Cell Metabolism ◽  
Mutant Cell ◽  
E3 Ubiquitin Ligase ◽  
Mapk Signaling ◽  
Ras Signaling ◽  
Growth Factor Signaling ◽  
N Terminus

AbstractDespite the prevalence of N-terminal acetylation (Nt-acetylation), little is known of its biological functions. In this study, we show that NatB regulates Rb mutant cell survival, EGFR/MAPK signaling activity, and EGFR signaling-dependent tumor growth. We identify Grb2/Drk, MAPK, and PP2AC as the key NatB targets of EGFR pathway. Surprisingly, NatB activity increases the levels of positive pathway components Grb2/Drk and MAPK while decreases the levels of negative pathway component PP2AC despite these proteins have the same first two amino acids that are recognized by NatB and N-end rule pathways. Mechanistically, we show that NatB regulates Grb2/Drk protein stability through its N-terminal sequences and that Grb2/Drk and MAPK are selectively degraded by the Arg/N-end rule E3 ubiquitin ligase Ubr4, which targets proteins with free N-terminus. In contrast, PP2AC is selectively degraded by the Ac/N-end rule pathway E3 ubiquitin ligase Cnot4 that targets proteins with acetylated N-terminus. These results reveal a novel mechanism by which NatB-mediated Nt-acetylation and N-end rule pathways modulate EGFR/MAPK signaling by inversely regulating the levels of positive and negative components. Since mutation or overexpression that deregulate the EGFR/Ras signaling pathway are common in human cancers and NatB subunits are significant unfavorable prognostic markers, this study can potentially lead to the development of novel therapeutic approaches.Significance StatementNt-acetylation is often regarded as a constitutive, irreversible, and static modification that is not suited to serve regulatory functions. Our observation that Nt-acetylation by NatB coordinately regulate the levels of positive and negative components of the EGFR/MAPK pathway show that Nt-acetylation and N-end rule pathways can play important roles regulating important signaling pathways. As Acetyl-CoA level, which is influenced by cell metabolism, can be rate limiting for Nt-acetylation, our results also suggest a potentially new mechanism by which cellular metabolic status can regulate growth factor signaling.

scholarly journals Putative E3 Ubiquitin Ligase of Human Rotavirus Inhibits NF-κB Activation by Using Molecular Mimicry To Target β-TrCP

mBio ◽  
2015 ◽  
Vol 6 (1) ◽  
Author(s):  
Marco Morelli ◽  
Allison F. Dennis ◽  
John T. Patton
Keyword(s):  
Ubiquitin Ligase ◽  
Molecular Mimicry ◽  
Nonstructural Protein ◽  
Critical Role ◽  
E3 Ubiquitin Ligase ◽  
Viral Proteins ◽  
Biochemical Properties ◽  
Antagonist Activity ◽  
Rotavirus A ◽  
Human Rotavirus

ABSTRACT  NF-κB plays a critical role in the induction and maintenance of innate and adaptive immune transcriptional programs. An associated inhibitor of κB protein (IκB) regulates NF-κB activation and contains a degron motif (DSGΦxS) that undergoes phosphorylation following pathogen recognition or other proinflammatory signals. The E3 ubiquitin ligase SCFβ-TrCPrecognizes this phosphodegron through its β-transducin repeat-containing protein (β-TrCP) subunit and induces IκB degradation, allowing NF-κB to translocate to the nucleus and modulate gene expression. Rotavirus (RV), a major cause of pediatric gastroenteritis, can block NF-κB activation through the action of its nonstructural protein NSP1, a putative E3 ubiquitin ligase that mediates the degradation of β-TrCP or other immunomodulatory proteins in a virus strain-specific manner. Here, we show that NSP1 targets β-TrCP by mimicking the IκB phosphodegron. The NSP1 proteins of most human and porcine RV strains conserve a C-terminal phosphodegron-like (PDL) motif, DSGΦS. Deletion of this motif or mutation of its serine residues disrupts NSP1-mediated degradation of β-TrCP and inhibition of NF-κB activation. Additionally, a point mutation within the phosphodegron-binding pocket protects β-TrCP from NSP1-mediated turnover. Fusion of the PDL motif to an NSP1 protein known to target other immunomodulatory proteins generates a chimeric NSP1 protein that can induce β-TrCP degradation and block NF-κB activation. Other viral proteins (Epstein-Barr virus LMP1, HIV-1 Vpu, and vaccinia virus A49) also contain a PDL motif and interact with β-TrCP to inhibit NF-κB activation. Taken together, these data suggest that targeting β-TrCP by molecular mimicry may be a common strategy used by human viruses to evade the host immune response.IMPORTANCE   The transcription factor NF-κB, a central regulator of the host response to infection, is a frequent target of viral antagonism. Pathogen detection activates NF-κB by inducing the phosphorylation of an associated inhibitor protein (IκB), which targets IκB for degradation by the E3 ubiquitin ligase β-TrCP. Rotavirus, a significant cause of childhood gastroenteritis, antagonizes NF-κB through the activity of its NSP1 protein, a putative E3 ubiquitin ligase that mediates β-TrCP turnover. Here, we show that NSP1 functions by mimicking the IκB phosphodegron recognized by β-TrCP. Nearly all human rotavirus strains conserve this motif at the NSP1 C terminus, and its removal disrupts NSP1 antagonist activity. This sequence conserves the biochemical properties of the IκB phosphodegron and can rescue antagonist activity when fused to an NSP1 protein otherwise inactive against β-TrCP. Other viral proteins also mimic IκB to disrupt NF-κB activation, indicating that this is an important immune evasion strategy.

scholarly journals The N Terminus of the Vaccinia Virus Protein F1L Is an Intrinsically Unstructured Region That Is Not Involved in Apoptosis Regulation

2016 ◽  
Vol 291 (28) ◽  
pp. 14600-14608 ◽  
Author(s):  
Sofia Caria ◽  
Bevan Marshall ◽  
Robyn-Lee Burton ◽  
Stephanie Campbell ◽  
Delara Pantaki-Eimany ◽  
...  
Keyword(s):  
Vaccinia Virus ◽  
Virus Protein ◽  
N Terminus ◽  
Apoptosis Regulation

scholarly journals Molecular basis of cullin-3 (Cul3) ubiquitin ligase subversion by vaccinia virus protein A55

2019 ◽  
Vol 294 (16) ◽  
pp. 6416-6429 ◽  
Author(s):  
Chen Gao ◽  
Mitchell A. Pallett ◽  
Tristan I. Croll ◽  
Geoffrey L. Smith ◽  
Stephen C. Graham
Keyword(s):  
Vaccinia Virus ◽  
Ubiquitin Ligase ◽  
Molecular Basis ◽  
Virus Protein ◽  
Cullin 3

scholarly journals Interplay between Polo kinase, LKB1-activated NUAK1 kinase, PP1βMYPT1 phosphatase complex and the SCFβTrCP E3 ubiquitin ligase

2014 ◽  
Vol 461 (2) ◽  
pp. 233-245 ◽  
Author(s):  
Sourav Banerjee ◽  
Anna Zagórska ◽  
Maria Deak ◽  
David G. Campbell ◽  
Alan R. Prescott ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Ubiquitin Ligase ◽  
E3 Ubiquitin Ligase ◽  
Biological Regulation ◽  
Polo Kinase ◽  
Suppress Cell Proliferation

The present study provides insights into the biological regulation of the NUAK isoforms and highlights the remarkable interplay that exists between Polo kinase, NUAK1, PP1βMYPT1 and SCFβTrCP signalling components. It demonstrates NUAK1 inhibitors suppress cell proliferation and PLK1.

scholarly journals Aurora B-dependent Ndc80 Degradation Regulates Kinetochore Composition in Meiosis

2019 ◽  
Author(s):  
Jingxun Chen ◽  
Andrew Liao ◽  
Emily N Powers ◽  
Hanna Liao ◽  
Lori A Kohlstaedt ◽  
...  
Keyword(s):  
Chromosome Segregation ◽  
Ubiquitin Ligase ◽  
Meiotic Prophase ◽  
Aurora B ◽  
Phosphorylation Sites ◽  
Independent Manner ◽  
N Terminus ◽  
Spindle Microtubules ◽  
Necessary And Sufficient ◽  
Single Subunit

ABSTRACTThe kinetochore complex is a conserved machinery that connects chromosomes to spindle microtubules. During meiosis, the kinetochore is restructured to accommodate a specialized chromosome segregation pattern. In budding yeast, meiotic kinetochore remodeling is mediated by the temporal changes in the abundance of a single subunit called Ndc80. We have previously described the regulatory events that control the timely synthesis of Ndc80. Here, we report that Ndc80 turnover is also tightly regulated in meiosis: Ndc80 degradation is active in meiotic prophase, but not in metaphase I. Ndc80 degradation depends on the ubiquitin ligase APCAma1 and is mediated by the proteasome. Importantly, Aurora B-dependent Ndc80 phosphorylation, a mark that has been previously implicated in correcting erroneous microtubule–kinetochore attachments, is essential for Ndc80 degradation in a microtubule-independent manner. The N-terminus of Ndc80, including a 27-residue sequence and Aurora B phosphorylation sites, is both necessary and sufficient for kinetochore protein degradation. Finally, defects in Ndc80 turnover predispose meiotic cells to chromosome mis-segregation. Our study elucidates the mechanism by which meiotic cells modulate their kinetochore composition through regulated Ndc80 degradation, and demonstrates that Aurora B-dependent regulation of kinetochores extends beyond altering microtubule attachments.

scholarly journals The SCFHOS/β-TRCP-ROC1 E3 Ubiquitin Ligase Utilizes Two Distinct Domains within CUL1 for Substrate Targeting and Ubiquitin Ligation

2000 ◽  
Vol 20 (4) ◽  
pp. 1382-1393 ◽  
Author(s):  
Kenneth Wu ◽  
Serge Y. Fuchs ◽  
Angus Chen ◽  
Peilin Tan ◽  
Carlos Gomez ◽  
...  
Keyword(s):  
Ubiquitin Ligase ◽  
Binding Activity ◽  
Dual Function ◽  
Necrosis Factor Alpha ◽  
Polyubiquitin Chains ◽  
C Terminus ◽  
N Terminus ◽  
Factor Alpha ◽  
F Box Protein ◽  
Necessary And Sufficient

ABSTRACT We describe a purified ubiquitination system capable of rapidly catalyzing the covalent linkage of polyubiquitin chains onto a model substrate, phosphorylated IκBα. The initial ubiquitin transfer and subsequent polymerization steps of this reaction require the coordinated action of Cdc34 and the SCFHOS/β-TRCP-ROC1 E3 ligase complex, comprised of four subunits (Skp1, cullin 1 [CUL1], HOS/β-TRCP, and ROC1). Deletion analysis reveals that the N terminus of CUL1 is both necessary and sufficient for binding Skp1 but is devoid of ROC1-binding activity and, hence, is inactive in catalyzing ubiquitin ligation. Consistent with this, introduction of the N-terminal CUL1 polypeptide into cells blocks the tumor necrosis factor alpha-induced and SCF-mediated degradation of IκB by forming catalytically inactive complexes lacking ROC1. In contrast, the C terminus of CUL1 alone interacts with ROC1 through a region containing the cullin consensus domain, to form a complex fully active in supporting ubiquitin polymerization. These results suggest the mode of action of SCF-ROC1, where CUL1 serves as a dual-function molecule that recruits an F-box protein for substrate targeting through Skp1 at its N terminus, while the C terminus of CUL1 binds ROC1 to assemble a core ubiquitin ligase.

scholarly journals Interaction between RING1 (R1) and the Ubiquitin-like (UBL) Domains Is Critical for the Regulation of Parkin Activity

2015 ◽  
Vol 291 (4) ◽  
pp. 1803-1816 ◽  
Author(s):  
Su Jin Ham ◽  
Soo Young Lee ◽  
Saera Song ◽  
Ju-Ryung Chung ◽  
Sekyu Choi ◽  
...  
Keyword(s):  
Ubiquitin Ligase ◽  
E3 Ubiquitin Ligase ◽  
E3 Ligase ◽  
Mitochondrial Outer Membrane ◽  
Missense Mutations ◽  
Mitochondrial Translocation ◽  
C Terminus ◽  
Carbonyl Cyanide ◽  
N Terminus ◽  
Upstream Kinase

Parkin is an E3 ligase that contains a ubiquitin-like (UBL) domain in the N terminus and an R1-in-between-ring-RING2 motif in the C terminus. We showed that the UBL domain specifically interacts with the R1 domain and negatively regulates Parkin E3 ligase activity, Parkin-dependent mitophagy, and Parkin translocation to the mitochondria. The binding between the UBL domain and the R1 domain was suppressed by carbonyl cyanide m-chlorophenyl hydrazone treatment or by expression of PTEN-induced putative kinase 1 (PINK1), an upstream kinase that phosphorylates Parkin at the Ser-65 residue of the UBL domain. Moreover, we demonstrated that phosphorylation of the UBL domain at Ser-65 prevents its binding to the R1 domain and promotes Parkin activities. We further showed that mitochondrial translocation of Parkin, which depends on phosphorylation at Ser-65, and interaction between the R1 domain and a mitochondrial outer membrane protein, VDAC1, are suppressed by binding of the UBL domain to the R1 domain. Interestingly, Parkin with missense mutations associated with Parkinson disease (PD) in the UBL domain, such as K27N, R33Q, and A46P, did not translocate to the mitochondria and induce E3 ligase activity by m-chlorophenyl hydrazone treatment, which correlated with the interaction between the R1 domain and the UBL domain with those PD mutations. These findings provide a molecular mechanism of how Parkin recruitment to the mitochondria and Parkin activation as an E3 ubiquitin ligase are regulated by PINK1 and explain the previously unknown mechanism of how Parkin mutations in the UBL domain cause PD pathogenesis.

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