Proteome-wide analysis of USP14 substrates revealed its role in hepatosteatosis via stabilization of FASN

Liu, Bin; Jiang, Shangwen; Li, Min; Xiong, Xuelian; Zhu, Mingrui; Li, Duanzhuo; Zhao, Lei; Qian, Lili; Zhai, Linhui; Li, Jing; Han, Lu; Sun, Shengnan; Lin, Jiandie D.; Lu, Yan; Li, Xiaoying; Tan, Minjia

doi:10.1038/s41467-018-07185-y

Cited by 92 publications

(86 citation statements)

References 48 publications

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“…A recent study, combining USP14 interactome and quantitative proteomics in response to USP14 knockdown, identified fatty acid synthase (FASN) as a specific USP14 substrate, and suggested a list of proteins as potential USP14 substrates (Liu et al, 2018). All these studies including ours start to reveal a picture of USP14 substrates.…”

Section: Discussionmentioning

confidence: 63%

Inactive USP14 and inactive UCHL5 cause accumulation of distinct ubiquitinated proteins in mammalian cells

Chadchankar

Korboukh

Doig

et al. 2018

Preprint

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6BT, UK (S.J.M.) Running title: Effects of inactive USP14 and UCHL5 in mammalian cellsAbstract USP14 is a cysteine-protease deubiquitinase associated with the proteasome and plays important catalytic and allosteric roles in proteasomal degradation. USP14 inhibition has been considered a therapeutic strategy for accelerating degradation of aggregation-prone proteins in neurodegenerative diseases and for inhibiting proteasome function to induce apoptotic cell death in cancers. Here we studied the effects of USP14 inhibition in mammalian cells using small molecule inhibitors and an inactive USP14 mutant C114A. Neither the inhibitors nor USP14 C114A changed the level of TDP-43, tau or α-synuclein in HEK293T cells. However, USP14 C114A led to an accumulation of ubiquitinated proteins, which were isolated by ubiquitin immunoprecipitation and identified by mass spectrometry. Among these proteins we confirmed that ubiquitinated β-catenin was accumulated in the cells expressing USP14 C114A with biochemistry and molecular biology experiments. The proteasome binding of USP14 C114A is required for its effect on ubiquitinated proteins. UCHL5 is the other cysteine-protease deubiquitinase associated with the proteasome. Interestingly, the inactive mutant of UCHL5 C88A also caused an accumulation of ubiquitinated proteins in HEK293T cells but did not affect β-catenin. Using ubiquitin immunoprecipitation and mass spectrometry, we identified the accumulated ubiquitinated proteins in UCHL5 C88A expressing cells which are mostly distinct from those accumulated in USP14 C114A expressing cells. Among the identified proteins are well established proteasome substrates and proteasome subunits. Together our data suggest that USP14 and UCHL5 can deubiquitinate distinct substrates at the proteasome and regulate the ubiquitination of the proteasome itself which is tightly linked to its function.

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Section: Discussionmentioning

confidence: 63%

Inactive USP14 and inactive UCHL5 cause accumulation of distinct ubiquitinated proteins in mammalian cells

Chadchankar

Korboukh

Doig

et al. 2018

Preprint

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“…The protein turnover and stability in eukaryotic cells is maintained by the ubiquitination/deubiquitination cascade (49). Our recent study found that aberrantly high levels of USP14 in the liver of obese mice and humans promote hepatosteatosis and hypertriglyceridemia by stabilizing fatty acid synthase (FASN) (32), which might increase the risk of glucose intolerance and T2DM (50). However, we found that FASN knockdown in the MPHs did not affect the gluconeogenic role of USP14 (SI Appendix, Fig.…”

Section: Discussionmentioning

confidence: 87%

“…The top 20 up-and downregulated genes are listed in SI Appendix, Fig. S3 B and C, and include the gene encoding for USP14, which has been shown to regulate hepatic triglyceride metabolism by our recent study (32). In addition, USP14 has been previously identified as a binding partner of IRE1α, and thereby a potential mediator of the UPR (33,34).…”

Section: Resultsmentioning

confidence: 98%

Sustained ER stress promotes hyperglycemia by increasing glucagon action through the deubiquitinating enzyme USP14

Liu

Zhang

et al. 2019

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

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Endoplasmic reticulum (ER) stress plays an important role in metabolic diseases like obesity and type 2 diabetes mellitus (T2DM), although the underlying mechanisms and regulatory pathways remain to be elucidated. Here, we induced chronic low-grade ER stress in lean mice to levels similar to those in high-fat diet (HFD)–fed obese mice and found that it promoted hyperglycemia due to enhanced hepatic gluconeogenesis. Mechanistically, sustained ER stress up-regulated the deubiquitinating enzyme ubiquitin-specific peptidase 14 (USP14), which increased the stability and levels of 3′,5′-cyclic monophosphate–responsive element binding (CREB) protein (CBP) to enhance glucagon action and hepatic gluconeogenesis. Exogenous overexpression of USP14 in the liver significantly increased hepatic glucose output. Consistent with this, liver-specific knockdown of USP14 abrogated the effects of ER stress on glucose metabolism, and also improved hyperglycemia and glucose intolerance in obese mice. In conclusion, our findings show a mechanism underlying ER stress-induced disruption of glucose homeostasis, and present USP14 as a potential therapeutic target against T2DM.

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“…In the present study, eliminating toxic proteins, facilitating DNA repair and other signaling pathways regulated by USP14 may also have accounted for the observed cisplatin insensitivity. Although many studies have demonstrated the importance of USP14 in cell physiology and diseases, the global substrates of USP14 are still to be elucidated, representing a major 'bottleneck' to uncover the functional characterization of USP14 and understand the complexity of proteasome-associated deubiquitination events (65,66).…”

Section: Discussionmentioning

confidence: 99%

Inhibition of ubiquitin‑specific protease�14 promotes connexin�32 internalization and counteracts cisplatin cytotoxicity in human ovarian cancer cells

Luo

Wang

et al. 2019

Oncol Rep

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Although cisplatin is one of the most accepted therapies for ovarian cancer, recurrence and drug resistance remain problematic. Both the ubiquitin-proteasome system (UPS) and connexin (Cx) are closely related to tumor progression. However, the role of ubiquitin-specific protease 14 (USP14) and Cx in mediating drug resistance remains unclear. In the present study, we aimed to determine whether USP14 is involved in cisplatin resistance and modulates the internalization of connexin 32 (Cx32) in ovarian cancer. The results of the deubiquitinase (DUB) trap assay and western blot analysis revealed that the expression and activity levels of USP14 were downregulated in A2780 cisplatin-resistant cells (A2780-CDDP) relative to these levels in A2780 cisplatin-sensitive cells (A2780). CCK-8 assay results showed that inhibition of USP14 by a specific inhibitor or siRNA decreased cisplatin cytotoxicity in A2780 cells. Additionally, USP14 inhibition increased the expression of Cx32 without changing its mRNA and ubiquitination levels, as showed by Real-time qPCR and immunoprecipitation assay respectively. Cisplatin resistance induced by USP14 inhibition was counteracted by Cx32 knockdown. Moreover, USP14 inhibition contributed to Cx32 internalization, as determined by western blot analysis and a reduction in gap junction intercellular communication (GJIC), as showed by parachute dye-coupling assay. Collectively, these data suggest that Cx32 internalization by USP14 inhibition modulates the cisplatin resistance in ovarian cancer cells, thus serving as a potential drug target to challenge chemotherapy failure. In addition, USP14 can also be used as a marker to monitor the development of cisplatin resistance in ovarian cancer treatment.

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Proteome-wide analysis of USP14 substrates revealed its role in hepatosteatosis via stabilization of FASN

Cited by 92 publications

References 48 publications

Inactive USP14 and inactive UCHL5 cause accumulation of distinct ubiquitinated proteins in mammalian cells

Inactive USP14 and inactive UCHL5 cause accumulation of distinct ubiquitinated proteins in mammalian cells

Sustained ER stress promotes hyperglycemia by increasing glucagon action through the deubiquitinating enzyme USP14

Inhibition of ubiquitin‑specific protease�14 promotes connexin�32 internalization and counteracts cisplatin cytotoxicity in human ovarian cancer cells

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