Role of Nse1 Subunit of SMC5/6 Complex as a Ubiquitin Ligase

Structural Maintenance of Chromosomes (SMC) complexes are important for many aspects of the chromosomal organization. Unlike cohesin and condensin, the SMC5/6 complex contains a variant RING domain carried by its Nse1 subunit. RING domains are characteristic for ubiquitin ligases, and human NSE1 has been shown to possess ubiquitin-ligase activity in vitro. However, other studies were unable to show such activity. Here, we confirm Nse1 ubiquitin-ligase activity using purified Schizosaccharomyces pombe proteins. We demonstrate that the Nse1 ligase activity is stimulated by Nse3 and Nse4. We show that Nse1 specifically utilizes Ubc13/Mms2 E2 enzyme and interacts directly with ubiquitin. We identify the Nse1 mutation (R188E) that specifically disrupts its E3 activity and demonstrate that the Nse1-dependent ubiquitination is particularly important under replication stress. Moreover, we determine Nse4 (lysine K181) as the first known SMC5/6-associated Nse1 substrate. Interestingly, abolition of Nse4 modification at K181 leads to suppression of DNA-damage sensitivity of other SMC5/6 mutants. Altogether, this study brings new evidence for Nse1 ubiquitin ligase activity, significantly advancing our understanding of this enigmatic SMC5/6 function.

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The Role of E3, E4 Ubiquitin Ligase (UBE4B) in Human Pathologies

Cancers ◽

10.3390/cancers12010062 ◽

2019 ◽

Vol 12 (1) ◽

pp. 62 ◽

Cited By ~ 1

Author(s):

Nikolaos Antoniou ◽

Nefeli Lagopati ◽

Dimitrios Ilias Balourdas ◽

Michail Nikolaou ◽

Alexandros Papalampros ◽

...

Keyword(s):

Dna Damage ◽

Dna Damage Response ◽

Ubiquitin Ligase ◽

Molecular Mechanisms ◽

Ubiquitin Ligases ◽

Cellular Functions ◽

P53 Expression ◽

Damage Response ◽

Mechanisms Of Carcinogenesis

The genome is exposed daily to many deleterious factors. Ubiquitination is a mechanism that regulates several crucial cellular functions, allowing cells to react upon various stimuli in order to preserve their homeostasis. Ubiquitin ligases act as specific regulators and actively participate among others in the DNA damage response (DDR) network. UBE4B is a newly identified member of E3 ubiquitin ligases that appears to be overexpressed in several human neoplasms. The aim of this review is to provide insights into the role of UBE4B ubiquitin ligase in DDR and its association with p53 expression, shedding light particularly on the molecular mechanisms of carcinogenesis.

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

Journal of Virology ◽

10.1128/jvi.01438-19 ◽

2019 ◽

Vol 93 (23) ◽

Cited By ~ 4

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.

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Liganded ERα Stimulates the E3 Ubiquitin Ligase Activity of UBE3C to Facilitate Cell Proliferation

Molecular Endocrinology ◽

10.1210/me.2015-1125 ◽

2015 ◽

Vol 29 (11) ◽

pp. 1646-1657 ◽

Cited By ~ 5

Author(s):

Maiko Okada ◽

Fumiaki Ohtake ◽

Hiroyuki Nishikawa ◽

Wenwen Wu ◽

Yasushi Saeki ◽

...

Keyword(s):

Cell Proliferation ◽

Ubiquitin Ligase ◽

E3 Ubiquitin Ligase ◽

Cyclin B1 ◽

Dependent Manner ◽

Ubiquitin Ligase Activity ◽

Dependent Cell ◽

Estrogen Antagonist

Abstract Estrogen receptor (ER)α is a well-characterized ligand-dependent transcription factor. However, the global picture of its nongenomic functions remains to be illustrated. Here, we demonstrate a novel function of ERα during mitosis that facilitates estrogen-dependent cell proliferation. An E3 ubiquitin ligase, UBE3C, was identified in an ERα complex from estrogen-treated MCF-7 breast cancer cells arrested at mitosis. UBE3C interacts with ERα during mitosis in an estrogen-dependent manner. In vitro, estrogen dramatically stimulates the E3 activity of UBE3C in the presence of ERα. This effect was inhibited by the estrogen antagonist tamoxifen. Importantly, estrogen enhances the ubiquitination of cyclin B1 (CCNB1) and destabilizes CCNB1 during mitosis in a manner dependent on endogenous UBE3C. ERα, UBE3C, and CCNB1 colocalize in prophase nuclei and at metaphase spindles before CCNB1 is degraded in anaphase. Depletion of UBE3C attenuates estrogen-dependent cell proliferation without affecting the transactivation function of ERα. Collectively, these results demonstrate a novel ligand-dependent action of ERα that stimulates the activity of an E3 ligase. The mitotic role of estrogen may contribute to its effects on proliferation in addition to its roles in target gene expression.

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Faculty Opinions recommendation of The ubiquitin ligase activity in the DDB2 and CSA complexes is differentially regulated by the COP9 signalosome in response to DNA damage.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1013426.192151 ◽

2003 ◽

Author(s):

Antony Carr

Keyword(s):

Dna Damage ◽

Ubiquitin Ligase ◽

Cop9 Signalosome ◽

Ubiquitin Ligase Activity

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LINC-PINT impedes DNA repair and enhances radiotherapeutic response by targeting DNA-PKcs in nasopharyngeal cancer

Cell Death and Disease ◽

10.1038/s41419-021-03728-2 ◽

2021 ◽

Vol 12 (5) ◽

Author(s):

You-hong Wang ◽

Zhen Guo ◽

Liang An ◽

Yong Zhou ◽

Heng Xu ◽

...

Keyword(s):

Dna Damage ◽

Nasopharyngeal Cancer ◽

Noncoding Rnas ◽

Dependent Protein Kinase ◽

Damage Repair ◽

Dependent Protein ◽

Cancer Tissues

AbstractRadioresistance continues to be the leading cause of recurrence and metastasis in nasopharyngeal cancer. Long noncoding RNAs are emerging as regulators of DNA damage and radioresistance. LINC-PINT was originally identified as a tumor suppressor in various cancers. In this study, LINC-PINT was significantly downregulated in nasopharyngeal cancer tissues than in rhinitis tissues, and low LINC-PINT expressions showed poorer prognosis in patients who received radiotherapy. We further identified a functional role of LINC-PINT in inhibiting the malignant phenotypes and sensitizing cancer cells to irradiation in vitro and in vivo. Mechanistically, LINC-PINT was responsive to DNA damage, inhibiting DNA damage repair through ATM/ATR-Chk1/Chk2 signaling pathways. Moreover, LINC-PINT increased radiosensitivity by interacting with DNA-dependent protein kinase catalytic subunit (DNA-PKcs) and negatively regulated the expression and recruitment of DNA-PKcs. Therefore, these findings collectively support the possibility that LINC-PINT serves as an attractive target to overcome radioresistance in NPC.

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The role of Schizosaccharomyces pombe Rad32, the Mre11 homologue, and other DNA damage response proteins in non-homologous end joining and telomere length maintenance

Nucleic Acids Research ◽

10.1093/nar/27.13.2655 ◽

1999 ◽

Vol 27 (13) ◽

pp. 2655-2661 ◽

Cited By ~ 50

Author(s):

S. Wilson ◽

N. Warr ◽

D. L. Taylor ◽

F. Z. Watts

Keyword(s):

Dna Damage ◽

Schizosaccharomyces Pombe ◽

Telomere Length ◽

Dna Damage Response ◽

End Joining ◽

Damage Response ◽

Non Homologous End Joining

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Genotoxicity and inflammatory potential of stainless steel welding fume particles: an in vitro study on standard vs Cr(VI)-reduced flux-cored wires and the role of released metals

Archives of Toxicology ◽

10.1007/s00204-021-03116-x ◽

2021 ◽

Author(s):

Sarah McCarrick ◽

Valentin Romanovski ◽

Zheng Wei ◽

Elin M. Westin ◽

Kjell-Arne Persson ◽

...

Keyword(s):

Dna Damage ◽

In Vitro Study ◽

Human Monocyte ◽

Welding Fumes ◽

Welding Fume ◽

Increased Risk ◽

Underlying Mechanisms ◽

Cored Wires

AbstractWelders are daily exposed to various levels of welding fumes containing several metals. This exposure can lead to an increased risk for different health effects which serves as a driving force to develop new methods that generate less toxic fumes. The aim of this study was to explore the role of released metals for welding particle-induced toxicity and to test the hypothesis that a reduction of Cr(VI) in welding fumes results in less toxicity by comparing the welding fume particles of optimized Cr(VI)-reduced flux-cored wires (FCWs) to standard FCWs. The welding particles were thoroughly characterized, and toxicity (cell viability, DNA damage and inflammation) was assessed following exposure to welding particles as well as their released metal fraction using cultured human bronchial epithelial cells (HBEC-3kt, 5–100 µg/mL) and human monocyte-derived macrophages (THP-1, 10–50 µg/mL). The results showed that all Cr was released as Cr(VI) for welding particles generated using standard FCWs whereas only minor levels (< 3% of total Cr) were released from the newly developed FCWs. Furthermore, the new FCWs were considerably less cytotoxic and did not cause any DNA damage in the doses tested. For the standard FCWs, the Cr(VI) released in cell media seemed to explain a large part of the cytotoxicity and DNA damage. In contrast, all particles caused rather similar inflammatory effects suggesting different underlying mechanisms. Taken together, this study suggests a potential benefit of substituting standard FCWs with Cr(VI)-reduced wires to achieve less toxic welding fumes and thus reduced risks for welders.

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Cellular Samurai: katanin and the severing of microtubules

Journal of Cell Science ◽

10.1242/jcs.113.16.2821 ◽

2000 ◽

Vol 113 (16) ◽

pp. 2821-2827 ◽

Cited By ~ 5

Author(s):

L. Quarmby

Keyword(s):

Epithelial Cells ◽

Essential Role ◽

Aaa Atpase ◽

C Elegans ◽

Microtubule Severing ◽

Biochemical Studies ◽

New Evidence

Recent biochemical studies of the AAA ATPase, katanin, provide a foundation for understanding how microtubules might be severed along their length. These in vitro studies are complemented by a series of recent reports of direct in vivo observation of microtubule breakage, which indicate that the in vitro phenomenon of catalysed microtubule severing is likely to be physiological. There is also new evidence that microtubule severing by katanin is important for the production of non-centrosomal microtubules in cells such as neurons and epithelial cells. Although it has been difficult to establish the role of katanin in mitosis, new genetic evidence indicates that a katanin-like protein, MEI-1, plays an essential role in meiosis in C. elegans. Finally, new proteins involved in the severing of axonemal microtubules have been discovered in the deflagellation system of Chlamydomonas.

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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.

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