Insights into degradation mechanism of N-end rule substrates by p62/SQSTM1 autophagy adapter

Kwon, Do Hoon; Park, Ok Hyun; Kim, Leehyeon; Jung, Yang Ouk; Park, Yeonkyoung; Jeong, Hyomin; Hyun, Jaekyung; Kim, Yoon Ki; Song, Hyun Kyu

doi:10.1038/s41467-018-05825-x

Cited by 62 publications

(73 citation statements)

References 61 publications

(72 reference statements)

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“…Lysine has a positive charge at physiological pH (7.4). The URB box and ZZ-domain of SQSTM1 utilize their negatively charged pocket to recognize and bind with positively charged protein-type 1 N-end rule substrates, including lysine, to promote protein lysosomal degradation [39]. We observed that the K222 site is completely exposed and extremely close to the N-terminus of LDHA (Fig.…”

Section: Discussionmentioning

confidence: 81%

Lysine-222 succinylation reduces lysosomal degradation of lactate dehydrogenase a and is increased in gastric cancer

Li¹,

Zhang²,

Zhao³

et al. 2020

J Exp Clin Cancer Res

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Background: Lysine succinylation is an emerging posttranslational modification that has garnered increased attention recently, but its role in gastric cancer (GC) remains underexplored. Methods: Proteomic quantification of lysine succinylation was performed in human GC tissues and adjacent normal tissues by mass spectrometry. The mRNA and protein levels of lactate dehydrogenase A (LDHA) in GC and adjacent normal tissues were analyzed by qRT-PCR and western blot, respectively. The expression of K222-succinylated LDHA was measured in GC tissue microarray by the K222 succinylation-specific antibody. The interaction between LDHA and sequestosome 1 (SQSTM1) was measured by co-immunoprecipitation (co-IP) and proximity ligation assay (PLA). The binding of carnitine palmitoyltransferase 1A (CPT1A) to LDHA was determined by co-IP. The effect of K222succinylated LDHA on tumor growth and metastasis was evaluated by in vitro and in vivo experiments. Results: Altogether, 503 lysine succinylation sites in 303 proteins were identified. Lactate dehydrogenase A (LDHA), the key enzyme in Warburg effect, was found highly succinylated at K222 in GC. Intriguingly, this modification did not affect LDHA ubiquitination, but reduced the binding of ubiquitinated LDHA to SQSTM1, thereby decreasing its lysosomal degradation. We demonstrated that CPT1A functions as a lysine succinyltransferase that interacts with and succinylates LDHA. Moreover, high K222-succinylation of LDHA was associated with poor prognosis in patients with GC. Finally, overexpression of a succinylation-mimic mutant of LDHA promoted cell proliferation, invasion, and migration. Conclusions: Our data revealed a novel lysosomal pathway of LDHA degradation, which is mediated by the binding of K63-ubiquitinated LDHA to SQSTM1. Strikingly, CPT1A succinylates LDHA on K222, which thereby reduces the binding and inhibits the degradation of LDHA, as well as promotes GC invasion and proliferation. This study thus uncovers a new role of lysine succinylation and the mechanism underlying LDHA upregulation in GC.

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Section: Discussionmentioning

confidence: 81%

Lysine-222 succinylation reduces lysosomal degradation of lactate dehydrogenase a and is increased in gastric cancer

Li¹,

Zhang²,

Zhao³

et al. 2020

J Exp Clin Cancer Res

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“…1B) (2,24,(66)(67)(68)). Yet another N-recognin of the human Arg/N-degron pathway is p62, an autophagyregulating protein distinct from E3s (41,69,70). hsUBR1 and hsUBR2 E3s are sequelogous (similar in sequence) (71) (they Fig.…”

Section: Significancementioning

confidence: 99%

Five enzymes of the Arg/N-degron pathway form a targeting complex: The concept of superchanneling

Hyun

Chen

et al. 2020

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

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The Arg/N-degron pathway targets proteins for degradation by recognizing their N-terminal (Nt) residues. If a substrate bears, for example, Nt-Asn, its targeting involves deamidation of Nt-Asn, arginylation of resulting Nt-Asp, binding of resulting (conjugated) Nt-Arg to the UBR1-RAD6 E3-E2 ubiquitin ligase, ligase-mediated synthesis of a substrate-linked polyubiquitin chain, its capture by the proteasome, and substrate’s degradation. We discovered that the human Nt-Asn–specific Nt-amidase NTAN1, Nt-Gln–specific Nt-amidase NTAQ1, arginyltransferase ATE1, and the ubiquitin ligase UBR1-UBE2A/B (or UBR2-UBE2A/B) form a complex in which NTAN1 Nt-amidase binds to NTAQ1, ATE1, and UBR1/UBR2. In addition, NTAQ1 Nt-amidase and ATE1 arginyltransferase also bind to UBR1/UBR2. In the yeast Saccharomyces cerevisiae, the Nt-amidase, arginyltransferase, and the double-E3 ubiquitin ligase UBR1-RAD6/UFD4-UBC4/5 are shown to form an analogous targeting complex. These complexes may enable substrate channeling, in which a substrate bearing, for example, Nt-Asn, would be captured by a complex-bound Nt-amidase, followed by sequential Nt modifications of the substrate and its polyubiquitylation at an internal Lys residue without substrate’s dissociation into the bulk solution. At least in yeast, the UBR1/UFD4 ubiquitin ligase interacts with the 26S proteasome, suggesting an even larger Arg/N-degron–targeting complex that contains the proteasome as well. In addition, specific features of protein-sized Arg/N-degron substrates, including their partly sequential and partly nonsequential enzymatic modifications, led us to a verifiable concept termed “superchanneling.” In superchanneling, the synthesis of a substrate-linked poly-Ub chain can occur not only after a substrate’s sequential Nt modifications, but also before them, through a skipping of either some or all of these modifications within a targeting complex.

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“…SQSTM1/p62 can bind to cargo molecules, such as misfolded proteins and their aggregates. Following binding, they are encapsulated together into the autophagosome and ultimately degraded by the lysosome in a suicide-like manner [56,57]. However, SQSTM1/p62 levels were increased and lysosomal activity was inhibited in leukemia cells following NTS exposure ( Figure 1F).…”

Section: Lysosome Inhibition Is Associated With Nts-induced Leukemiamentioning

confidence: 99%

Non-Thermal Plasma Induces Antileukemic Effect Through mTOR Ubiquitination

Kim

et al. 2020

Cells

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Non-thermal plasma (NTP) has been studied as a novel therapeutic tool for cancer that does not damage healthy cells. In this study, we show that NTP-treated solutions (NTS) can induce death in various leukemia cells through mechanistic target of rapamycin (mTOR) ubiquitination. Previously, we manufactured and demonstrated the efficacy of NTS in solid cancers. NTS did not exhibit any deleterious side effects, such as acute death or weight loss in nude mice. In the present study, NTS induced cell death in myeloid leukemia cells, including acute myeloid leukemia (AML) and chronic myeloid leukemia (CML). We found that mTOR was downregulated in NTS-treated cells via the ubiquitin-proteasome system (UPS). We also identified ‘really interesting new gene’ finger protein 126 (RNF126) as a novel binding protein for mTOR through protein arrays and determined the role of E3 ligase in NTS-induced mTOR ubiquitination. NTS-derived reactive oxygen species (ROS) affected RNF126 expression and lysosomal dysfunction. These findings suggest that NTS has potential antileukemic effects through RNF126-mediated mTOR ubiquitination with no deleterious side effects. Thus, NTS may represent a new therapeutic method for chemotherapy-resistant leukemia.

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Insights into degradation mechanism of N-end rule substrates by p62/SQSTM1 autophagy adapter

Cited by 62 publications

References 61 publications

Lysine-222 succinylation reduces lysosomal degradation of lactate dehydrogenase a and is increased in gastric cancer

Lysine-222 succinylation reduces lysosomal degradation of lactate dehydrogenase a and is increased in gastric cancer

Five enzymes of the Arg/N-degron pathway form a targeting complex: The concept of superchanneling

Non-Thermal Plasma Induces Antileukemic Effect Through mTOR Ubiquitination

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