Direct involvement of the ubiquitin-conjugating enzyme Ubc9/Hus5 in the degradation of IκBα

Tashiro, Kei; Pando, Matthew P.; Kanegae, Yumi; Wamsley, Penny; Inoue, Satoshi; Verma, Inder M.

doi:10.1073/pnas.94.15.7862

Cited by 49 publications

(53 citation statements)

References 40 publications

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“…3 It is also well established that regulators of NF-kB, the Ikba proteins are degraded rapidly via the proteasomal pathway during inflammation leading to enhanced shuttling of NF-kB to the nucleus favoring cytokine gene expression. 4 Cells do have potent mechanisms for stabilizing Ikba such as UBC9-mediated SUMO-1 conjugation of this protein that prevents its degradation. 10 However, inflammatory signal buildup in cells can overcome this stabilization effect.…”

Section: Discussionmentioning

confidence: 99%

“…3 The NF-kB proteins play a major role in LPS-mediated inflammatory response by activating the expression of the above cytokine genes. 4 The activity of NF-kB is regulated by its interaction with the NF-kB inhibitor a protein (Ikba) that is known to prevent nuclear translocation of NF-kB, retaining it in the cytoplasm. 4 Activation of Kupffer cells appears to modulate liver injury 5 and is also known to trigger the induction of inflammatory cytokines in hepatocytes.…”

mentioning

confidence: 99%

“…4 The activity of NF-kB is regulated by its interaction with the NF-kB inhibitor a protein (Ikba) that is known to prevent nuclear translocation of NF-kB, retaining it in the cytoplasm. 4 Activation of Kupffer cells appears to modulate liver injury 5 and is also known to trigger the induction of inflammatory cytokines in hepatocytes. 5 Hepatocytes respond to LPS-mediated stimulation by inducing the levels of inducible nitric oxide synthase (iNOS).…”

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confidence: 99%

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Ubiquitin-Conjugating Enzyme 9 Phosphorylation as a Novel Mechanism for Potentiation of the Inflammatory Response

Tomasi

Ramani

Ryoo

2016

The American Journal of Pathology

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Lipopolysaccharide (LPS), a bacterial endotoxin, induces inflammation in macrophages via activation of NF-kB signaling. Sumoylation is a post-translational modification mediated by the small ubiquitin-like modifier, SUMO. Ubiquitin-conjugating enzyme 9 (UBC9) is the only known SUMO conjugating enzyme. LPS treatment lowers SUMO-1 and UBC9 mRNA levels in primary astrocytes. UBC9 can degrade NF-kB inhibitor a (Ikba) via a SUMO2/3-ubiquitin pathway. However, UBC9 may also promote Ikba stability by SUMO-1 conjugation that further regulates NF-kB signaling. The role of UBC9 in liver inflammation is unknown. We reported that CDK1-mediated phosphorylation of UBC9 enhanced its stability. Herein, we describe an anti-inflammatory role of UBC9 that is lost when it is phosphorylated during inflammation. LPS exposure caused induction in UBC9 phosphorylation and CDK1 activation specifically in Kupffer cells in vivo and in RAW264.7 macrophages in vitro. Silencing or overexpression experiments in vitro and in vivo showed that UBC9 was required to blunt the proinflammatory response elicited by LPS. LPS stimulation raised the binding of phospho-UBC9 but not the unphosphorylated counterpart, to Ikba in RAW264.7 macrophages. Hence, phospho-UBC9 may promote NF-kB signaling by regulating Ikba and this may be a novel mechanism that deregulates liver inflammatory signaling. (Am J Pathol 2016 http://dx

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Section: Discussionmentioning

confidence: 99%

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confidence: 99%

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confidence: 99%

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Ubiquitin-Conjugating Enzyme 9 Phosphorylation as a Novel Mechanism for Potentiation of the Inflammatory Response

Tomasi

Ramani

Ryoo

2016

The American Journal of Pathology

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“…Dominantnegative inhibitors with mutations of this residue have been used to analyse the function of many E2s (Banerjee et al, 1995;Tashiro et al, 1997;Townsley et al, 1997;Pati et al, 1999;Gu and Roizman, 2003;Li et al, 2003). We used site-directed mutagenesis to create such a mutant molecule, UBE2Q2 (C304A) , referred to as DN-UBE2Q2-FLAG.…”

Section: Dominant-negative Ube2q2 Potentiates the Mitotic Arrest Indumentioning

confidence: 99%

Inactivation of the ubiquitin conjugating enzyme UBE2Q2 causes a prophase arrest and enhanced apoptosis in response to microtubule inhibiting agents

2007

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A putative ubiquitin conjugating enzyme known as UBE2Q2 was previously identified in a microarray screen for mitotic regulatory proteins. UBE2Q2 is very similar to another human protein, UBE2Q1 and orthologs from other higher eukaryotic species. In these studies, we demonstrate that UBE2Q2 can covalently bind ubiquitin on the active site cysteine in vitro and show that inhibition of this protein in vivo causes an early mitotic arrest and increased cytotoxicity when cells are treated with microtubule inhibiting agents (MIAs). Changes in cell cycle progression and viability are not observed in the absence of MIA treatment, indicating that UBE2Q2 is involved in the response to MIAs rather than performing a more general function in mitosis. Inhibition of the UBE2Q2 protein causes cells to undergo a prolonged prophase arrest suggesting that UBE2Q2 normally functions to antagonize an early mitotic checkpoint. Furthermore, UBE2Q2 inhibition sensitizes cells to the cytotoxic effects of MIAs through caspase-mediated apoptosis that is correlated with PARP-1 cleavage. These data provide insights into the cellular response to MIAs and demonstrate that inhibition of UBE2Q2 protein function may be useful in the treatment of malignancies.

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“…2A (compare lanes 1 and 2), CPT treatment of JN362a expressing hTOP1 (no exogenous Smt3p) resulted in the formation of high molecular weight hTOP1 species (see bands marked with * ) as revealed by immunoblotting with anti-hTOP1 antibodies. When His-6͞FLAG-tagged Smt3p was 25 M CPT in 1% DMSO (ϩ) or 1% DMSO (Ϫ) for 15 min and then lysed using the alkali lysis procedure described in Materials and Methods. Samples were analyzed on 6% SDS-PAGE and immunoblotted with anti-hTOP1 antibodies.…”

Section: Cpt Induces Rapid Covalent Modification Of Htop1 In Both Hummentioning

confidence: 99%

SUMO-1 conjugation to topoisomerase I: A possible repair response to topoisomerase-mediated DNA damage

Mao¹,

Sun²,

Desai³

et al. 2000

Proc. Natl. Acad. Sci. U.S.A.

187

148

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Ubiquitin͞26S proteasome-dependent degradation of topoisomerase I (TOP1) has been suggested to be a unique repair response to TOP1-mediated DNA damage. In the current study, we show that treatment of mammalian cells or yeast cells expressing human DNA TOP1 with camptothecin (CPT) induces covalent modification of the TOP1 by SUMO-1͞Smt3p, a ubiquitin-like protein. This conclusion is based on the following observations: (i) Mammalian DNA TOP1 conjugates induced by CPT were cross-reactive with SUMO-1͞Smt3p-specific antibodies both in yeast expressing human DNA TOP1 as well as mammalian cells. (ii) The formation of TOP1 conjugates was shown to be dependent on UBC9, the E2 enzyme for SUMO-1͞Smt3p. (iii) TOP1 physically interacts with UBC9. (iv) Ubc9 mutant yeast cells expressing human DNA TOP1 was hypersensitive to CPT, suggesting that UBC9͞SUMO-1 may be involved in the repair of TOP1-mediated DNA damage.UBC9 ͉ Smt3p ͉ ubiquitin T opoisomerase-mediated DNA damage represents an unique type of DNA damage whose importance has become increasingly appreciated. Many antibiotics, anticancer drugs, toxins, carcinogens, and physiological stresses are known to abort the catalytic cycles of topoisomerases resulting in the formation of topoisomerase-mediated DNA damage (1-7). Despite its importance, little is known about the molecular basis for the repair of this unique type of DNA damage.Recent studies have demonstrated that topoisomerase I-(TOP1) mediated DNA damage activates a ubiquitin͞proteasome pathway, resulting in degradation of TOP1 (down-regulation of TOP1) (8). It has been speculated that this pathway may represent a unique repair mechanism for TOP1-mediated DNA damage because down-regulation of human TOP1 (hTOP1) is expected to result in resistance͞tolerance to TOP1 poisons (8).Human SUMO-1 (small ubiquitin-like modifier) is an 11-kDa protein that shares 18% sequence homology with ubiquitin (9-11). Smt3p, which is 48% identical to human SUMO-1, has been identified in yeast (12,13). SUMO-1͞Smt3p is activated by a heterodimeric E1 enzyme in both the yeast and mammalian systems (13)(14)(15). In addition, UBC9, an E2 enzyme, has been shown to specifically conjugate SUMO-1͞Smt3p to the target proteins (11,(16)(17)(18). Recently, an enzyme Ulp1p, which specifically cleaves proteins from SUMO-1͞Smt3p conjugates and is distinct from isopeptidases for ubiquitin conjugates, also has been identified in yeast (19). These results suggest that the SUMO-1͞Smt3p pathway is very similar to the ubiquitin pathway but distinct E1 and E2 enzymes as well as proteases are involved in these pathways. The biological function of the SUMO-1͞Smt3p pathway appears diverse. UBC9 was originally identified to play an essential function in G 2 -M cell cycle progression in yeast (20). However, UBC9 is also known to be important for DNA repair (21). It has been shown that SUMO-1 can be conjugated to a number of proteins such as PML (22), RanGAP1 (23, 24), I B␣ (25), FAS͞apolipoprotein-1 (26), p53 (27, 28), RAD51, and RAD52 (29). A growing number of pr...

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Direct involvement of the ubiquitin-conjugating enzyme Ubc9/Hus5 in the degradation of IκBα

Cited by 49 publications

References 40 publications

Ubiquitin-Conjugating Enzyme 9 Phosphorylation as a Novel Mechanism for Potentiation of the Inflammatory Response

Ubiquitin-Conjugating Enzyme 9 Phosphorylation as a Novel Mechanism for Potentiation of the Inflammatory Response

Inactivation of the ubiquitin conjugating enzyme UBE2Q2 causes a prophase arrest and enhanced apoptosis in response to microtubule inhibiting agents

SUMO-1 conjugation to topoisomerase I: A possible repair response to topoisomerase-mediated DNA damage

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