scholarly journals Molecular mechanisms suppressing COP1 / SPA E3 ubiquitin ligase activity in blue light

2020 ◽  
Vol 169 (3) ◽  
pp. 418-429 ◽  
Author(s):  
Jathish Ponnu
Keyword(s):  
Blue Light ◽  
Ubiquitin Ligase ◽  
Molecular Mechanisms ◽  
E3 Ubiquitin Ligase ◽  
Ubiquitin Ligase Activity

scholarly journals The E3 Ubiquitin Ligase TBK1 Mediates the Degradation of Multiple Picornavirus VP3 Proteins by Phosphorylation and Ubiquitination

2019 ◽  
Vol 93 (23) ◽  
Author(s):  
Dan Li ◽  
Wenping Yang ◽  
Jingjing Ren ◽  
Yi Ru ◽  
Keshan Zhang ◽  
...  
Keyword(s):  
Ubiquitin Ligase ◽  
Molecular Mechanisms ◽  
E3 Ubiquitin Ligase ◽  
Adaptor Protein ◽  
Ubiquitin Ligase Activity ◽  
Lysine Residues ◽  
Innate Antiviral Immunity

ABSTRACT TANK-binding kinase 1 (TBK1) is essential for interferon beta (IFN-β) production and innate antiviral immunity. However, other, additional functions of TBK1 have remained elusive. Here, we showed that TBK1 is an E3 ubiquitin ligase that undergoes self-ubiquitylation in vitro in the presence of the E2 enzyme UbcH5c. Further evidence showed that TBK1 could also be self-ubiquitylated in vivo. Importantly, multiple picornavirus VP3 proteins were degraded by TBK1 through its kinase and E3 ubiquitin ligase activity. Mechanistically, TBK1 phosphorylated multiple picornavirus VP3 proteins at serine residues and ubiquitinated them via K63-linked ubiquitination at lysine residues. In addition, the C426 and C605 residues of TBK1 were not essential for TBK1 innate immunity activity; however, these residues were required for degradation of multiple picornavirus VP3 proteins and for its E3 ubiquitin ligase activity. Hence, our findings identified a novel role of TBK1 in regulating the virus life cycle and provided new insights into the molecular mechanisms of TBK1-mediated antiviral response. IMPORTANCE TBK1 is an important adaptor protein required for innate immune response to viruses, but its other functions were unknown. In this study, we found that TBK1 is an E3 ubiquitin ligase that undergoes self-ubiquitylation in vitro in the presence of the E2 enzyme UbcH5c. In addition, multiple picornavirus VP3 proteins were degraded by TBK1 through its kinase and E3 ubiquitin ligase activity. Our report provides evidence that TBK1 plays a role in viral protein degradation.

scholarly journals Allosteric Interactions byp53mRNA Govern HDM2 E3 Ubiquitin Ligase Specificity under Different Conditions

2016 ◽  
Vol 36 (16) ◽  
pp. 2195-2205 ◽  
Author(s):  
Ixaura Medina-Medina ◽  
Paola García-Beltrán ◽  
Ignacio de la Mora-de la Mora ◽  
Jesús Oria-Hernández ◽  
Guy Millot ◽  
...  
Keyword(s):  
Dna Damage ◽  
Substrate Specificity ◽  
Ubiquitin Ligase ◽  
Molecular Mechanisms ◽  
E3 Ubiquitin Ligase ◽  
Intrinsically Disordered ◽  
Allosteric Interactions ◽  
Ubiquitin Ligase Activity ◽  
Positive Regulators ◽  
Ring Interaction

HDM2 and HDMX are key negative regulatory factors of the p53 tumor suppressor under normal conditions by promoting its degradation or preventing itstransactivity, respectively. It has more recently been shown that both proteins can also act as positive regulators of p53 after DNA damage. This involves phosphorylation by ATM on serine residues HDM2(S395) and HDMX(S403), promoting their respective interaction with thep53mRNA. However, the underlying molecular mechanisms of how these phosphorylation events switch HDM2 and HDMX from negative to positive regulators of p53 is not known. Our results show that these phosphorylation events reside within intrinsically disordered domains and change the conformation of the proteins. The modifications promote the exposition of N-terminal interfaces that support the formation of a new HDMX-HDM2 heterodimer independent of the C-terminal RING-RING interaction. The E3 ubiquitin ligase activity of this complex toward p53 is prevented by thep53mRNA ligand but, interestingly, does not affect the capacity to ubiquitinate HDMX and HDM2. These results show how ATM-mediated modifications of HDMX and HDM2 switch HDM2 E3 ubiquitin ligase activity away from p53 but toward HDMX and itself and illustrate how the substrate specificity of HDM2 E3 ligase activity is regulated.

scholarly journals Id1 enhances RING1b E3 ubiquitin ligase activity through the Mel-18/Bmi-1 polycomb group complex

Oncogene ◽  
2010 ◽  
Vol 29 (43) ◽  
pp. 5818-5827 ◽  
Author(s):  
T Qian ◽  
J-Y Lee ◽  
J-H Park ◽  
H-J Kim ◽  
G Kong
Keyword(s):  
Ubiquitin Ligase ◽  
E3 Ubiquitin Ligase ◽  
Polycomb Group ◽  
Ubiquitin Ligase Activity

Post-translational regulation of FLC is mediated by an E3 ubiquitin ligase activity of SINAT5 in Arabidopsis

Plant Science ◽  
2007 ◽  
Vol 173 (2) ◽  
pp. 269-275 ◽  
Author(s):  
Bong Soo Park ◽  
Wan Gyu Sang ◽  
Song Yion Yeu ◽  
Yang Do Choi ◽  
Nam-Chon Paek ◽  
...  
Keyword(s):  
Ubiquitin Ligase ◽  
Translational Regulation ◽  
E3 Ubiquitin Ligase ◽  
Ubiquitin Ligase Activity

scholarly journals Autographa californica Nucleopolyhedrovirus AC141 (Exon0), a Potential E3 Ubiquitin Ligase, Interacts with Viral Ubiquitin and AC66 To Facilitate Nucleocapsid Egress

2017 ◽  
Vol 92 (3) ◽  
Author(s):  
Siddhartha Biswas ◽  
Leslie G. Willis ◽  
Minggang Fang ◽  
Yingchao Nie ◽  
David A. Theilmann
Keyword(s):  
Ubiquitin Ligase ◽  
Nuclear Export ◽  
Molecular Mechanisms ◽  
Nuclear Periphery ◽  
E3 Ubiquitin Ligase ◽  
Autographa Californica ◽  
Nucleocapsid Proteins ◽  
Systemic Spread ◽  
Autographa Californica Nucleopolyhedrovirus ◽  
Budded Virus

ABSTRACTDuring the infection cycle of Autographa californica multiple nucleopolyhedrovirus (AcMNPV), two forms of virions are produced, budded virus (BV) and occlusion-derived virus (ODV). Nucleocapsids that form BV have to egress from the nucleus, whereas nucleocapsids that form ODV remain inside the nucleus. The molecular mechanism that determines whether nucleocapsids remain inside or egress from the nucleus is unknown. AC141 (a predicted E3 ubiquitin ligase) and viral ubiquitin (vUbi) have both been shown to be required for efficient BV production. In this study, it was hypothesized that vUbi interacts with AC141, and in addition, that this interaction was required for BV production. Deletion of bothac141andvubirestricted viral infection to a single cell, and BV production was completely eliminated. AC141 was ubiquitinated by either vUbi or cellular Ubi, and this interaction was required for optimal BV production. Nucleocapsids in BV, but not ODV, were shown to be specifically ubiquitinated by vUbi, including a 100-kDa protein, as well as high-molecular-weight conjugates. The viral ubiquitinated 100-kDa BV-specific nucleocapsid protein was identified as AC66, which is known to be required for BV production and was shown by coimmunoprecipitation and mass spectrometry to interact with AC141. Confocal microscopy also showed that AC141, AC66, and vUbi interact at the nuclear periphery. These results suggest that ubiquitination of nucleocapsid proteins by vUbi functions as a signal to determine if a nucleocapsid will egress from the nucleus and form BV or remain in the nucleus to form ODV.IMPORTANCEBaculoviruses produce two types of virions called occlusion-derived virus (ODV) and budded virus (BV). ODVs are required for oral infection, whereas BV enables the systemic spread of virus to all host tissues, which is critical for killing insects. One of the important steps for BV production is the export of nucleocapsids out of the nucleus. This study investigated the molecular mechanisms that enable the selection of nucleocapsids for nuclear export instead of being retained within the nucleus, where they would become ODV. Our data show that ubiquitination, a universal cellular process, specifically tags nucleocapsids of BV, but not those found in ODV, using a virus-encoded ubiquitin (vUbi). Therefore, ubiquitination may be the molecular signal that determines if a nucleocapsid is destined to form a BV, thus ensuring lethal infection of the host.

Functions of RMR proteins in intracellular trafficking in the moss "Physcomitrium patens" Mitten (previously "Physcomitrella patens")

2021 ◽  
Author(s):  
◽  
Carla Coppola
Keyword(s):  
Ubiquitin Ligase ◽  
Physcomitrella Patens ◽  
Fluorescent Protein ◽  
Storage Proteins ◽  
Higher Plants ◽  
E3 Ubiquitin Ligase ◽  
E3 Ligases ◽  
Ubiquitin Ligase Activity ◽  
Vacuolar Transport

In this study, I focused on a new family of receptors, called RMRs (Receptor-like Membrane RING-H2) and I tried to investigate their role in the moss Physcomitrium patens Mitten (previously Physcomitrella patens). There is some evidence that in Angiosperms, RMRs are vacuolar receptors for the neutral/storage vacuole that is a compartment where storage proteins and metabolites are accumulated during seeds development or in somatic tissues. It is distinguished from lytic vacuole which has the same functions as animal lysosomes. The five PpRMR genes have been knocked-out, yielding viable material without visible phenotype (Ayachi, 2012). A trafficking phenotype was described by Fahr (2017) who generated the construct Citrine-Cardosin (Ci-Card) composed of the fluorescent protein Citrine fused to the C-terminal vacuolar sorting determinant (ctVSD) from cardosin A (cardosin is addressed to the vacuole in higher plants —Pereira et al., 2013). The fusion protein was delivered to the central vacuole of PpWT but mistargeted in PpRMR-KO lines, indicating that the targeting of this protein to the vacuole depends on PpRMRs. The introduction of this thesis presents the plant endomembrane system, with particular attention to vacuolar transport and ubiquitylation. In the second chapter, I show the techniques used to attempt to detect PpRMRs by Western Blot: our failure may be due to a rapid degradation of these proteins, which could prevent their detection. In the third chapter, I focused on PpRMR2 involvement in ubiquitylation. We hypothesize that PpRMRs are E3 ligases because they are members of the PA-TM-RING protein family. Most of these proteins have an E3 ubiquitin ligase activity in animals (Seroogy et al., 2004; Borchers et al., 2002), for this reason, we think that plant PpRMRs could have this function as well, which could contribute to vacuolar targeting. Indeed, I could confirm that PpRMR2 has an E3 ubiquitin ligase activity. PpRMRs substrates are still unknown in moss thus we have analysed putative candidates supposing that they could be ubiquitylated by PpRMRs. We have tested this hypothesis through in vitro ubiquitylation assays, obtaining ambiguous results. In the fourth chapter, I show preliminary results about the visible phenotype of PpRMR-KO mutants: PpWT and PpRMR-KO lines displayed phenotypic differences in leafy gametophores, which were accentuated upon salt stress exposure. Lastly, I transformed the transgenic lines PpWT/Ci-Card and Pp5KO/Ci-Card with mutated versions of PpRMR2 and analysed their effect on vacuolar transport by confocal microscopy. For most of the constructions tested, the trafficking was perturbed in both lines. Only PpWT/Ci-Card expressing PpRMR2ΔSer (lacking the Serine-Rich motif) displayed a typical vacuolar pattern.

scholarly journals Identification of a PGXPP degron motif in dishevelled and structural basis for its binding to the E3 ligase KLHL12

Open Biology ◽  
2020 ◽  
Vol 10 (6) ◽  
pp. 200041 ◽  
Author(s):  
Zhuoyao Chen ◽  
Gregory A. Wasney ◽  
Sarah Picaud ◽  
Panagis Filippakopoulos ◽  
Masoud Vedadi ◽  
...  
Keyword(s):  
Ubiquitin Ligase ◽  
Molecular Mechanisms ◽  
Hydrophobic Interactions ◽  
E3 Ubiquitin Ligase ◽  
Structural Basis ◽  
E3 Ligases ◽  
Kelch Domain ◽  
Ring E3 Ligases ◽  
Ring E3 ◽  
New Protein

Wnt signalling is dependent on dishevelled proteins (DVL1-3), which assemble an intracellular Wnt signalosome at the plasma membrane. The levels of DVL1-3 are regulated by multiple Cullin-RING E3 ligases that mediate their ubiquitination and degradation. The BTB-Kelch protein KLHL12 was the first E3 ubiquitin ligase to be identified for DVL1-3, but the molecular mechanisms determining its substrate interactions have remained unknown. Here, we mapped the interaction of DVL1-3 to a ‘PGXPP' motif that is conserved in other known partners and substrates of KLHL12, including PLEKHA4, PEF1, SEC31 and DRD4. To determine the binding mechanism, we solved a 2.4 Å crystal structure of the Kelch domain of KLHL12 in complex with a DVL1 peptide that bound with low micromolar affinity. The DVL1 substrate adopted a U-shaped turn conformation that enabled hydrophobic interactions with all six blades of the Kelch domain β-propeller. In cells, the mutation or deletion of this motif reduced the binding and ubiquitination of DVL1 and increased its stability confirming this sequence as a degron motif for KLHL12 recruitment. These results define the molecular mechanisms determining DVL regulation by KLHL12 and establish the KLHL12 Kelch domain as a new protein interaction module for a novel proline-rich motif.

scholarly journals E3 ubiquitin ligase MARCHF5 controls BAK apoptotic activity independently of BH3-only proteins

2022 ◽  
Author(s):  
Grant Dewson ◽  
Alan Shuai Huang ◽  
Hui San Chin ◽  
Boris Reljic ◽  
Tirta M Djajawi ◽  
...  
Keyword(s):  
Ubiquitin Ligase ◽  
Apoptotic Cell ◽  
E3 Ubiquitin Ligase ◽  
Ubiquitin Ligase Activity ◽  
A Genome ◽  
Library Screen ◽  
Important Regulator ◽  
Apoptotic Activity ◽  
Bh3 Mimetic ◽  
Survival Proteins

Intrinsic apoptosis is principally governed by the BCL-2 family of proteins, but some non-BCL-2 proteins are also critical to control this process. To identify novel apoptosis regulators, we performed a genome-wide CRISPR-Cas9 library screen, and identified the mitochondrial E3 ubiquitin ligase MARCHF5/MITOL/RNF153 as an important regulator of BAK apoptotic function. Deleting MARCHF5 in diverse cell lines dependent on BAK conferred profound resistance to BH3-mimetic drugs. The loss of MARCHF5 or its E3 ubiquitin ligase activity surprisingly drove BAK to adopt an activated conformation, with resistance to BH3-mimetics afforded by the formation of inhibitory complexes with pro-survival proteins MCL-1 and BCL-XL. Importantly, these changes to BAK conformation and pro-survival association occurred independently of BH3-only proteins and influence on pro-survival proteins. This study identifies a new mechanism by which MARCHF5 regulates apoptotic cell death and provides new insight into how cancer cells respond to BH3-mimetic drugs. These data also highlight the emerging role of ubiquitin signalling in apoptosis that may be exploited therapeutically.

scholarly journals Inhibition of cIAP1 as a strategy for targeting c-MYC–driven oncogenic activity

2018 ◽  
Vol 115 (40) ◽  
pp. E9317-E9324 ◽  
Author(s):  
Haoyan Li ◽  
Yanjia Fang ◽  
Chunyi Niu ◽  
Hengyi Cao ◽  
Ting Mi ◽  
...  
Keyword(s):  
Ubiquitin Ligase ◽  
E3 Ubiquitin Ligase ◽  
E3 Ligase ◽  
Pharmacological Agents ◽  
Protein Levels ◽  
Ubiquitin Ligase Activity ◽  
Molecule Inhibitor ◽  
Smac Mimetics ◽  
High Throughput Assay ◽  
In Cells

Protooncogenec-MYC, a master transcription factor, is a major driver of human tumorigenesis. Development of pharmacological agents for inhibiting c-MYC as an anticancer therapy has been a longstanding but elusive goal in the cancer field. E3 ubiquitin ligase cIAP1 has been shown to mediate the activation of c-MYC by destabilizing MAD1, a key antagonist of c-MYC. Here we developed a high-throughput assay for cIAP1 ubiquitination and identified D19, a small-molecule inhibitor of E3 ligase activity of cIAP1. We show that D19 binds to the RING domain of cIAP1 and inhibits the E3 ligase activity of cIAP1 by interfering with the dynamics of its interaction with E2. Blocking cIAP1 with D19 antagonizes c-MYC by stabilizing MAD1 protein in cells. Furthermore, we show that D19 and an improved analog (D19-14) promote c-MYC degradation and inhibit the oncogenic function of c-MYC in cells and xenograft animal models. In contrast, we show that activating E3 ubiquitin ligase activity of cIAP1 by Smac mimetics destabilizes MAD1, the antagonist of MYC, and increases the protein levels of c-MYC. Our study provides an interesting example using chemical biological approaches for determining distinct biological consequences from inhibiting vs. activating an E3 ubiquitin ligase and suggests a potential broad therapeutic strategy for targeting c-MYC in cancer treatment by pharmacologically modulating cIAP1 E3 ligase activity.

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