Proteasome-Mediated Degradation of p21 via N-Terminal Ubiquitinylation

Bloom, Joanna; Amador, Virginia; Bartolini, Francesca; DeMartino, George N.; Pagano, Michele

doi:10.1016/s0092-8674(03)00755-4

Cited by 290 publications

(268 citation statements)

References 24 publications

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“…p21 protein can undergo ubiquitination‐mediated proteasomal degradation during cell cycle progression 32. Based on this notion, we treated DAOY and D283MED cells with proteasome inhibitor MG‐132 in time course experiments.…”

Section: Resultsmentioning

confidence: 99%

“…According to other studies reporting that IAPs function as E3 ligases for both ubiquitination and neddylation,32 we next examined whether LBW242 treatment‐increased p21 protein expression due to abrogation of NEDD8 (neddylation)‐mediated proteasomal degradation. MB cells were treated with NEDD8‐activating enzyme (NAE) inhibitor MLN4924 and their p21 protein levels were determined at different time points.…”

Section: Resultsmentioning

confidence: 99%

“…It is possible that IAPs participate in regulation of p21 protein stability. Additionally, p21 protein has been reported to undergo degradation via ubiquitin‐ or NEDD8‐proteasome system 32. Hence, we investigated whether p21 undergoes ubiquitin‐dependent proteasomal degradation.…”

Section: Discussionmentioning

confidence: 99%

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Targeting inhibitors of apoptosis proteins suppresses medulloblastoma cell proliferation via G2/M phase arrest and attenuated neddylation of p21

et al. 2018

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Medulloblastoma (MB) is the most common type of malignant childhood brain tumor. We previously showed that inhibitors of apoptosis proteins (IAP) small‐molecule inhibitors (LCL161 or LBW242) combined with chemotherapy have synergistic antiproliferative effects on MB cells. The synergistic antitumor effects of combination treatments happen through induction of autophagy and caspase‐3/7‐activated apoptosis. Here, we investigated the effects of IAP inhibitors or silencing IAP on cell cycle regulation. We discovered that treatment with IAP inhibitors or their combination with conventional chemotherapy (vincristine or cisplatin), as well as RNAi knockdown of cIAP1/2 or XIAP arrested MB cells in the G2/M phase through downregulation of cyclin B1‐CDK1 and cyclin A‐CDK1/2. Among these three IAPs, only silencing cIAP1 expression enhanced p21 dependent‐G2/M phase accumulation. IAP inhibitors reduced cIAP1 expression and increased p21 expression in time course experiments. Furthermore, cIAP1 can govern p21 proteasomal degradation via neddylation in lieu of ubiquitination. Inhibition of IAPs significantly abrogated cIAP1‐mediated p21 degradation. We also observed an inverse correlation between nuclear cIAP1 and nuclear p21 expressions in MB tumor tissues. These findings provide new mechanistic evidence of the influence of IAP inhibitors on MB cell proliferation through disruption of the cell cycle.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

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Targeting inhibitors of apoptosis proteins suppresses medulloblastoma cell proliferation via G2/M phase arrest and attenuated neddylation of p21

et al. 2018

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“…In addition, BubR1 knockdown via RNAi caused a drastic reduction of p21 in HeLa cells that do not have detectable levels of p53 before or after DNA damage (data not shown). As p21 is also degraded by the proteasome-mediated pathway (Bloom et al, 2003), it is interesting to know whether elevated APC/C Cdc20 activity would modulate its poly-ubiquitylation in BubR1-deficient cells. A recent study shows that APC/C Cdh1 participates in the regulation of SCF(Skp2-Cks1) activity, which in turn is responsible for the degradation of p21 and p27 (Bashir et al, 2004).…”

Section: Discussionmentioning

confidence: 99%

BubR1 is involved in regulation of DNA damage responses

Fang

Wang

et al. 2006

Oncogene

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“…Second, both linear and isopeptide fusions have physiological relevance. Ubiquitination at the amino termini of substrate proteins can occur in vivo and lead to target protein degradation [42][43][44]. In addition, DUBs are responsible for the normal processing and maturation of the genome-encoded ubiquitin precursors, all of which are linear fusion proteins.…”

mentioning

confidence: 99%

Small ubiquitin-like modifying protein isopeptidase assay based on poliovirus RNA polymerase activity

Arnold

Bernal

Uche

et al. 2006

Analytical Biochemistry

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The ubiquitin-proteasome pathway is the major nonlysosomal proteolytic system in eukaryotic cells responsible for regulating the level of many key regulatory molecules within the cells. Modification of cellular proteins by ubiquitin and ubiquitin-like proteins, such as small ubiquitin-like modifying protein (SUMO), plays an essential role in a number of biological schemes, and ubiquitin pathway enzymes have become important therapeutic targets. Ubiquitination is a dynamic reversible process; a multitude of ubiquitin ligases and deubiquitinases (DUBs) are responsible for the wide-ranging influence of this pathway as well as its selectivity. The DUB enzymes serve to maintain adequate pools of free ubiquitin and regulate the ubiquitination status of cellular proteins. Using SUMO fusions, a novel assay system, based on poliovirus RNA-dependent RNA polymerase activity, is described here. The method simplifies the isopeptidase assay and facilitates high-throughput analysis of these enzymes. The principle of the assay is the dependence of the viral polymerase on a free N terminus for activity; accordingly, the polymerase is inactive when fused at its N terminus to SUMO or any other ubiquitin-like protein. The assay is sensitive, reproducible, and adaptable to a highthroughput format for use in screens for inhibitors/activators of clinically relevant SUMO proteases and deubiquitinases. Keywords Isopeptidases; Protein degradation; N terminus; 3D polymeraseThe ubiquitin-proteasomal pathway regulates the cellular content and/or compartmentalization of many proteins [1,2] and is a promising, albeit underexploited, area for drug discovery. The therapeutic value of this pathway was recently validated by the introduction and clinical success of Velcade, a proteasome inhibitor, for the treatment of refractory relapsed multiple myeloma [3]. Other ubiquitin-like proteins (UBLs) 1 have recently been shown to contribute similarly to the cellular regulation of proteins, with the most extensively characterized UBL being small ubiquitin-like modifying protein (SUMO) [4].Destruction of a targeted protein via the ubiquitin system initially involves recognition by a multienzyme system that attaches ubiquitin to the target protein. Energy is required to activate ubiquitin at its carboxy terminus. Activation is catalyzed by the enzyme E1, which links the ubiquitin C terminus to a cysteine side chain of the enzyme. This activated ubiquitin is * Corresponding author. Fax: +1 610 644 8616., E-mail address: mattern@progenra.com (M.R. Mattern).. 1 Abbreviations used: UBL, ubiquitin-like protein; SUMO, small ubiquitin-like modifying protein; UBP/USP, ubiquitin-specific protease; UCH, ubiquitin terminal hydrolase; DUB, deubiquitinating enzyme; Ub-AMC, Ub-7-amino-4-methylcoumarin; EDTA, ethylenediaminetetraacetic acid; IPTG, isopropyl-β-D-thiogalactopyranoside; PMSF, phenylmethylsulfonylfluoride; GFP, green fluorescent protein; DTT, dithiothreitol. transferred to a second enzyme of the series, E2 (ubiquitin-conjugating protein), as a thioe...

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Proteasome-Mediated Degradation of p21 via N-Terminal Ubiquitinylation

Cited by 290 publications

References 24 publications

Targeting inhibitors of apoptosis proteins suppresses medulloblastoma cell proliferation via G2/M phase arrest and attenuated neddylation of p21

Targeting inhibitors of apoptosis proteins suppresses medulloblastoma cell proliferation via G2/M phase arrest and attenuated neddylation of p21

BubR1 is involved in regulation of DNA damage responses

Small ubiquitin-like modifying protein isopeptidase assay based on poliovirus RNA polymerase activity

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