AtPPRT1, an E3 Ubiquitin Ligase, Enhances the Thermotolerance in Arabidopsis

E3 ubiquitin ligase plays a vital role in the ubiquitin-mediated heat-related protein degradation pathway. Herein, we report that the expression of AtPPRT1, a C3HC4 zinc-finger ubiquitin E3 ligase gene, was induced by heat stress, and the β-glucuronidase (GUS) gene driven by the AtPPRT1 promoter has shown increased activity after basal and acquired thermotolerance. To further explore the function of AtPPRT1 in heat stress response (HSR), we used the atpprt1 mutant and AtPPRT1-overexpressing lines (OE2 and OE10) to expose in heat shock. In this study, the atpprt1 mutant had a lower germination and survival rate than those of Col-0 when suffered from the heat stress, whereas OEs enhanced basal and acquired thermotolerance in Arabidopsis seedlings. When compared to Col-0 and OEs, loss-of-function in AtPPRT1 resulted in lower chlorophyll retention and higher content of reactive oxygen species (ROS) after heat treatment. Moreover, the transcript levels of AtPPRT1 and several heat-related genes (AtZAT12, AtHSP21 and AtHSFA7a) were upregulated to greater extents in OEs and lower extents in atpprt1 compared to Col-0 after heat treated. Hence, we suggest that AtPPRT1 may act as a positive role in regulating the high temperature by mediating the degradation of unknown target proteins.

Download Full-text

Abstract 18: Synoviolin, an E3 Ubiquitin Ligase, Modulates Cardiac Myocyte Size and Restores Heart Function in Hypertrophic Cardiomyopathy

Circulation Research ◽

10.1161/res.111.suppl_1.a18 ◽

2012 ◽

Vol 111 (suppl_1) ◽

Author(s):

Shirin Doroudgar ◽

Mirko Völkers ◽

Donna J Thuerauf ◽

Ashley Bumbar ◽

Mohsin Khan ◽

...

Keyword(s):

Ubiquitin Ligase ◽

Protein Misfolding ◽

Cardiac Myocyte ◽

Protein Quality ◽

Heart Function ◽

E3 Ubiquitin Ligase ◽

Ubiquitin Ligases ◽

Calcium Handling ◽

Loss Of Function

The endoplasmic reticulum (ER) is essential for protein homeostasis, or proteostasis, which governs the balance of the proteome. In addition to secreted and membrane proteins, proteins bound for many other cellular locations are also made on ER-bound ribosomes, emphasizing the importance of protein quality and quantity control in the ER. Unlike cytosolic E3 ubiquitin ligases studied in the heart, synoviolin/Hrd1, which has not been studied in the heart, is an ER transmembrane E3 ubiquitin ligase, which we found to be upregulated upon protein misfolding in cardiac myocytes. Given the strategic location of synoviolin in the ER membrane, we addressed the hypothesis that synoviolin is critical for regulating the balance of the proteome, and accordingly, myocyte size. We showed that in vitro, adenovirus-mediated overexpression of synoviolin decreased cardiac myocyte size and protein synthesis, but unlike atrophy-related ubiquitin ligases, synoviolin did not increase global protein degradation. Furthermore, targeted gene therapy using adeno-associated virus 9 (AAV9) showed that overexpression of synoviolin in the left ventricle attenuated maladaptive cardiac hypertrophy and preserved cardiac function in mice subjected to trans-aortic constriction (AAV9-control TAC = 22.5 ± 6.2% decrease in EF vs. AAV9-synoviolin TAC at 6 weeks post TAC; P<0.001), and decreased mTOR activity. Since calcium is a major regulator of cardiac myocyte size, we examined the effects of synoviolin gain- or loss-of-function, using AAV9-synoviolin, or an miRNA designed to knock down synoviolin, respectively. While synoviolin gain-of-function did not affect calcium handling in isolated adult myocytes, synoviolin loss-of-function increased calcium transient amplitude (P<0.01), prolonged spark duration (P<0.001), and increased spark width (P<0.001). Spark frequency and amplitude were unaltered upon synoviolin gain- or loss-of-function. Whereas SR calcium load was unaltered by synoviolin loss-of-function, SERCA-mediated calcium removal was reduced (P<0.05). In conclusion, our studies suggest that in the heart, synoviolin is 1) a critical component of proteostasis, 2) a novel determinant of cardiac myocyte size, and 3) necessary for proper calcium handling.

Download Full-text

Cul4-Ddb1 ubiquitin ligases facilitate DNA replication-coupled sister chromatid cohesion through regulation of cohesin acetyltransferase Esco2

10.1101/414888 ◽

2018 ◽

Cited By ~ 1

Author(s):

Haitao Sun ◽

Jiaxin Zhang ◽

Jingjing Zhang ◽

Zhen Li ◽

Qinhong Cao ◽

...

Keyword(s):

Dna Replication ◽

Ubiquitin Ligase ◽

Sister Chromatid ◽

Critical Role ◽

E3 Ubiquitin Ligase ◽

Ubiquitin Ligases ◽

Sister Chromatid Cohesion ◽

Vital Role ◽

Chromatid Cohesion ◽

Evolutionarily Conserved

AbstractCohesin acetyltransferases Esco1 and Esco2 play a vital role in establishing sister chromatid cohesion. How Esco1 and Esco2 are controlled to achieve this in a DNA replication-coupled manner remains unclear in higher eukaryotes. Here we show that Cul4-RING ligases (CRL4s) play a critical role in sister chromatid cohesion in human cells. Depletion of Cul4A, Cul4B or Ddb1 subunits substantially reduces normal cohesion efficiency. We also show that Mms22L, a vertebrate ortholog of yeast Mms22, is one of Ddb1 and Cul4-associated factors (DCAFs) involved in cohesion. Several lines of evidence suggest a selective interaction of CRL4s with Esco2, but not Esco1. Depletion of either CRL4s or Esco2 causes a defect in Smc3 acetylation which can be rescued by HDAC8 inhibition. More importantly, both CRL4s and PCNA act as mediators for efficiently stabilizing Esco2 on chromatin and catalyzing Smc3 acetylation. Taken together, we propose an evolutionarily conserved mechanism in which CRL4s and PCNA regulate Esco2-dependent establishment of sister chromatid cohesion.Author summaryWe identified human Mms22L as a substrate specific adaptor of Cul4-Ddb1 E3 ubiquitin ligase. Downregulation of Cul4A, Cul4B or Ddb1 subunit causes reduction of acetylated Smc3, via interaction with Esco2 acetyltransferase, and then impairs sister chromatid cohesion in 293T cells. We found functional complementation between Cul4-Ddb1-Mms22L E3 ligase and Esco2 in Smc3 acetylation and sister chromatid cohesion. Interestingly, both Cul4-Ddb1 E3 ubiquitin ligase and PCNA contribute to Esco2 mediated Smc3 acetylation. To summarise, we demonstrated an evolutionarily conserved mechanism in which Cul4-Ddb1 E3 ubiquitin ligases and PCNA regulate Esco2-dependent establishment of sister chromatid cohesion.

Download Full-text

An E3 ubiquitin ligase fromNicotiana benthamianatargets the replicase ofBamboo mosaic virusand restricts its replication

10.1101/434076 ◽

2018 ◽

Author(s):

I-Hsuan Chen ◽

Jui-En Chang ◽

Chen-Yu Wu ◽

Ying-Ping Huang ◽

Yau-Huei Hsu ◽

...

Keyword(s):

Ubiquitin Ligase ◽

Fluorescent Protein ◽

Transmembrane Domain ◽

Tobacco Rattle Virus ◽

E3 Ubiquitin Ligase ◽

Infection Process ◽

Ring Domain ◽

Virus Induced Gene Silencing ◽

Ubiquitin E3 Ligase ◽

Rapid Amplification

AbstractOne upregulated host gene identified previously was found involved in the infection process ofBamboo mosaic virus(BaMV). The full-length cDNA of this gene was cloned by 5′- and 3′-rapid amplification of cDNA ends and found to encode a polypeptide containing a conserved RING-domain and a transmembrane domain. The gene might function as an E3 ubiquitin ligase. We designated this protein inNicotiana benthamianaas ubiquitin E3 ligase containing RING-domain 1 (NbUbE3R1). Further characterization by usingTobacco rattle virus-based virus-induced gene silencing revealed an increased BaMV accumulation in both knockdown plants and protoplasts. To further inspect the functional role of NbUbE3R1 in BaMV accumulation, NbUbE3R1 was expressed inN. benthamianaplants. The wild-type NbUbE3R1-orange fluorescent protein (NbUbE3R1-OFP), NbUbE3R1/△TM-OFP (removal of the transmembrane domain) and NbUbE3R1/mRING-OFP (mutation at the RING domain, the E2 interaction site) were transiently expressed in plants. NbUbE3R1 and its derivatives all functioned in restricting BaMV accumulation. The common feature of these constructs was the intact substrate-interacting domain. Yeast two-hybrid and co-immunoprecipitation experiments used to determine the possible viral-encoded substrate of NbUbE3R1 revealed the replicase of BaMV as the possible substrate. In conclusion, we identified an upregulated gene, NbUbE3R1, that plays a role in BaMV replication.

Download Full-text

PRMT5 epigenetically regulates the E3 ubiquitin ligase ITCH to influence lipid accumulation during mycobacterial infection

10.1101/2021.11.09.467864 ◽

2021 ◽

Author(s):

Salik Miskat Borbora ◽

R.S. Rajmani ◽

Kithiganahalli N Balaji

Keyword(s):

Ubiquitin Ligase ◽

Lipid Accumulation ◽

Ex Vivo ◽

E3 Ubiquitin Ligase ◽

Mycobacterial Infection ◽

Drug Regimen ◽

Loss Of Function ◽

Methyl Transferase ◽

Accumulation Of Lipids

Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis (TB), triggers enhanced accumulation of lipids to generate foamy macrophages (FMs). This process has been often attributed to the surge in the expression of lipid influx genes with a concomitant decrease in those involved in lipid efflux genes. Here, we define an Mtb-orchestrated modulation of the ubiquitination mechanism of lipid accumulation markers to enhance lipid accretion during infection. We find that Mtb infection represses the expression of the E3 ubiquitin ligase, ITCH, resulting in the sustenance of key lipid accrual molecules viz. ADRP and CD36, that are otherwise targeted by ITCH for proteasomal degradation. In line, overexpressing ITCH in Mtb-infected cells was found to suppress Mtb-induced lipid accumulation. Molecular analyses including loss-of-function and ChIP assays demonstrated a role for the concerted action of the transcription factor YY1 and the arginine methyl transferase PRMT5 in restricting the expression of Itch gene by conferring repressive symmetrical H4R3me2 marks on its promoter. Consequently, siRNA-mediated depletion of YY1 or PRMT5 rescued ITCH expression, thereby compromising the levels of Mtb-induced ADRP and CD36 and limiting FM formation during infection. Accumulation of lipids within the host has been implicated as a pro-mycobacterial process that aids in pathogen persistence and dormancy. In our study, perturbation of PRMT5 enzyme activity resulted in compromised lipid levels and reduced mycobacterial survival in primary murine macrophages (ex vivo) and in a therapeutic mouse model of TB infection (in vivo). These findings provide new insights on the role of PRMT5 and YY1 in augmenting mycobacterial pathogenesis. Thus, we posit that our observations could help design novel adjunct therapies and combinatorial drug regimen for effective anti-TB strategies.

Download Full-text

The E3 ubiquitin ligase RNF186 and RNF186 risk variants regulate innate receptor-induced outcomes

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2013500118 ◽

2021 ◽

Vol 118 (32) ◽

pp. e2013500118

Author(s):

Kishu Ranjan ◽

Matija Hedl ◽

Clara Abraham

Keyword(s):

Ubiquitin Ligase ◽

E3 Ubiquitin Ligase ◽

Bacterial Clearance ◽

Loss Of Function ◽

Complex Molecules ◽

Downstream Signaling ◽

Signaling Complex ◽

Risk Variants ◽

Ubiquitin Ligase Activity ◽

Inflammatory Bowel

Balancing microbial-induced cytokines and microbial clearance is critical at mucosal sites such as the intestine. How the inflammatory bowel disease (IBD)–associated gene RNF186 regulates this balance is unclear. We found that macrophages from IBD-risk rs6426833 carriers in the RNF186 region showed reduced cytokines to stimulation through multiple pattern recognition receptors (PRRs). Upon stimulation of PRRs, the E3-ubiquitin ligase RNF186 promoted ubiquitination of signaling complex molecules shared across PRRs and those unique to select PRRs. Furthermore, RNF186 was required for PRR-initiated signaling complex assembly and downstream signaling. RNF186, along with its intact E3-ubiquitin ligase activity, was required for optimal PRR-induced antimicrobial reactive oxygen species, reactive nitrogen species, and autophagy pathways and intracellular bacterial clearance in human macrophages and for bacterial clearance in intestinal myeloid cells. Cells transfected with the rare RNF186-A64T IBD-risk variant and macrophages from common rs6426833 RNF186 IBD-risk carriers demonstrated a reduction in these RNF186-dependent outcomes. These studies identify mechanisms through which RNF186 regulates innate immunity and show that RNF186 IBD-risk variants demonstrate a loss of function in PRR-initiated outcomes.

Download Full-text

The Arabidopsis E3 ubiquitin ligase PUB4 regulates BIK1 homeostasis and is targeted by a bacterial type-III effector

10.1101/2020.10.25.354514 ◽

2020 ◽

Author(s):

Maria Derkacheva ◽

Gang Yu ◽

Jose S. Rufian ◽

Shushu Jiang ◽

Paul Derbyshire ◽

...

Keyword(s):

Ubiquitin Ligase ◽

Plant Immunity ◽

E3 Ubiquitin Ligase ◽

Bacterial Pathogen ◽

Positive Role ◽

Type Iii ◽

Molecular Pattern ◽

Pathogen Associated Molecular Pattern ◽

Bacterial Type ◽

Shed Light

SummaryPlant immunity is tightly controlled by a complex and dynamic regulatory network, which ensures optimal activation upon detection of potential pathogens. Accordingly, each component of this network is a potential target for manipulation by pathogens. Here, we report that RipAC, a type III-secreted effector from the bacterial pathogen Ralstonia solanacearum, targets the plant E3 ubiquitin ligase PUB4 to inhibit pattern-triggered immunity (PTI). PUB4 plays a positive role in PTI by regulating the homeostasis of the central immune kinase BIK1. Before PAMP perception, PUB4 promotes the degradation of non-activated BIK1, while, after PAMP perception, PUB4 contributes to the accumulation of activated BIK1. RipAC leads to BIK1 degradation, which correlates with its PTI-inhibitory activity. RipAC causes a reduction in pathogen-associated molecular pattern (PAMP)-induced PUB4 accumulation and phosphorylation. Our results shed light on the role played by PUB4 in immune regulation, and illustrate an indirect targeting of the immune signalling hub BIK1 by a bacterial effector.

Download Full-text

AtPPRT3, a novel E3 ubiquitin ligase, plays a positive role in ABA signaling

Plant Cell Reports ◽

10.1007/s00299-020-02575-3 ◽

2020 ◽

Vol 39 (11) ◽

pp. 1467-1478 ◽

Cited By ~ 1

Author(s):

Yu Liu ◽

Lu Peng ◽

Xuemeng Gao ◽

Yingying Liu ◽

Zhibin Liu ◽

...

Keyword(s):

Ubiquitin Ligase ◽

E3 Ubiquitin Ligase ◽

Aba Signaling ◽

Positive Role

Download Full-text

Stringent requirement for HRD1, SEL1L, and OS-9/XTP3-B for disposal of ERAD-LS substrates

The Journal of Cell Biology ◽

10.1083/jcb.200910042 ◽

2010 ◽

Vol 188 (2) ◽

pp. 223-235 ◽

Cited By ~ 121

Author(s):

Riccardo Bernasconi ◽

Carmela Galli ◽

Verena Calanca ◽

Toshihiro Nakajima ◽

Maurizio Molinari

Keyword(s):

Ubiquitin Ligase ◽

E3 Ubiquitin Ligase ◽

Degradation Pathway ◽

Control Mechanisms ◽

Stringent Requirement ◽

Folding Intermediates ◽

Cytosolic Domains ◽

Membrane Changes ◽

Selection Of

Sophisticated quality control mechanisms prolong retention of protein-folding intermediates in the endoplasmic reticulum (ER) until maturation while sorting out terminally misfolded polypeptides for ER-associated degradation (ERAD). The presence of structural lesions in the luminal, transmembrane, or cytosolic domains determines the classification of misfolded polypeptides as ERAD-L, -M, or -C substrates and results in selection of distinct degradation pathways. In this study, we show that disposal of soluble (nontransmembrane) polypeptides with luminal lesions (ERAD-LS substrates) is strictly dependent on the E3 ubiquitin ligase HRD1, the associated cargo receptor SEL1L, and two interchangeable ERAD lectins, OS-9 and XTP3-B. These ERAD factors become dispensable for degradation of the same polypeptides when membrane tethered (ERAD-LM substrates). Our data reveal that, in contrast to budding yeast, tethering of mammalian ERAD-L substrates to the membrane changes selection of the degradation pathway.

Download Full-text

The E3 ubiquitin ligase SP1-like 1 plays a positive role in peroxisome biogenesis in Arabidopsis

The Plant Journal ◽

10.1111/tpj.13900 ◽

2018 ◽

Vol 94 (5) ◽

pp. 836-846 ◽

Cited By ~ 8

Author(s):

Ronghui Pan ◽

John Satkovich ◽

Cheng Chen ◽

Jianping Hu

Keyword(s):

Ubiquitin Ligase ◽

E3 Ubiquitin Ligase ◽

Peroxisome Biogenesis ◽

Positive Role

Download Full-text

A Noncanonical Mechanism of Nrf2 Activation by Autophagy Deficiency: Direct Interaction between Keap1 and p62

Molecular and Cellular Biology ◽

10.1128/mcb.00248-10 ◽

2010 ◽

Vol 30 (13) ◽

pp. 3275-3285 ◽

Cited By ~ 468

Author(s):

Alexandria Lau ◽

Xiao-Jun Wang ◽

Fei Zhao ◽

Nicole F. Villeneuve ◽

Tongde Wu ◽

...

Keyword(s):

Ubiquitin Ligase ◽

Defense Mechanism ◽

Ectopic Expression ◽

E3 Ubiquitin Ligase ◽

Direct Interaction ◽

Degradation Pathway ◽

26S Proteasome ◽

Ubiquitin Ligase Complex ◽

Subsequent Degradation ◽

Kelch Domain

ABSTRACT In response to stress, cells can utilize several cellular processes, such as autophagy, which is a bulk-lysosomal degradation pathway, to mitigate damages and increase the chances of cell survival. Deregulation of autophagy causes upregulation of p62 and the formation of p62-containing aggregates, which are associated with neurodegenerative diseases and cancer. The Nrf2-Keap1 pathway functions as a critical regulator of the cell's defense mechanism against oxidative stress by controlling the expression of many cellular protective proteins. Under basal conditions, Nrf2 is ubiquitinated by the Keap1-Cul3-E3 ubiquitin ligase complex and targeted to the 26S proteasome for degradation. Upon induction, the activity of the E3 ubiquitin ligase is inhibited through the modification of cysteine residues in Keap1, resulting in the stabilization and activation of Nrf2. In this current study, we identified the direct interaction between p62 and Keap1 and the residues required for the interaction have been mapped to 349-DPSTGE-354 in p62 and three arginines in the Kelch domain of Keap1. Accumulation of endogenous p62 or ectopic expression of p62 sequesters Keap1 into aggregates, resulting in the inhibition of Keap1-mediated Nrf2 ubiquitination and its subsequent degradation by the proteasome. In contrast, overexpression of mutated p62, which loses its ability to interact with Keap1, had no effect on Nrf2 stability, demonstrating that p62-mediated Nrf2 upregulation is Keap1 dependent. These findings demonstrate that autophagy deficiency activates the Nrf2 pathway in a noncanonical cysteine-independent mechanism.

Download Full-text