Potent USP10/13 antagonist spautin‐1 suppresses melanoma growth via ROS‐mediated DNA damage and exhibits synergy with cisplatin

Guo, Jia; Jiangling, Zhang; Liang, Long; Liu, Nian; Qi, Min; Zhao, Shuang; Su, Juan; Liu, Jing; Peng, Cong; Liu, Hong

doi:10.1111/jcmm.15093

Cited by 32 publications

(29 citation statements)

References 54 publications

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“…Autophagy inhibitors might work in later stages of infec-tion to dampen viral production, but this will depend on whether autophagy is active at the time or if the constituent components have been captured and effectively shut down. Several inhibitors/ activators have been reported (a selection is listed in Table 2), which can target autophagy and multiple auxiliary signals feeding into the process of autophagy (204)(205)(206)(207). One such axis is via the mTORC1…”

Section: Slims and Their Potential Therapeutic Implicationsmentioning

confidence: 99%

Short linear motif candidates in the cell entry system used by SARS-CoV-2 and their potential therapeutic implications

et al. 2021

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The first reported receptor for SARS-CoV-2 on host cells was the angiotensin-converting enzyme 2 (ACE2). However, the viral spike protein also has an RGD motif, suggesting that cell surface integrins may be co-receptors. We examined the sequences of ACE2 and integrins with the Eukaryotic Linear Motif (ELM) resource and identified candidate short linear motifs (SLiMs) in their short, unstructured, cytosolic tails with potential roles in endocytosis, membrane dynamics, autophagy, cytoskeleton, and cell signaling. These SLiM candidates are highly conserved in vertebrates and may interact with the μ2 subunit of the endocytosis-associated AP2 adaptor complex, as well as with various protein domains (namely, I-BAR, LC3, PDZ, PTB, and SH2) found in human signaling and regulatory proteins. Several motifs overlap in the tail sequences, suggesting that they may act as molecular switches, such as in response to tyrosine phosphorylation status. Candidate LC3-interacting region (LIR) motifs are present in the tails of integrin β3 and ACE2, suggesting that these proteins could directly recruit autophagy components. Our findings identify several molecular links and testable hypotheses that could uncover mechanisms of SARS-CoV-2 attachment, entry, and replication against which it may be possible to develop host-directed therapies that dampen viral infection and disease progression. Several of these SLiMs have now been validated to mediate the predicted peptide interactions.

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Section: Slims and Their Potential Therapeutic Implicationsmentioning

confidence: 99%

Short linear motif candidates in the cell entry system used by SARS-CoV-2 and their potential therapeutic implications

et al. 2021

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“…Because both shUSP7 and shUSP10 have puromycin resistance, we could not generate a stable cell line with concomitant knockdown of both DUBs. Instead, we used HBX41108 and spautin‐1 to inhibit USP7 and USP10, respectively 24,25 . Treating Caco‐2bbe/NHE3 with HBX41108 or spoutin‐1 decreased NHE3 protein abundance by 26.8 ± 2.2% and 14.5 ± 2.4%, respectively, whereas a 43.7 ± 1.9% reduction was achieved with both inhibitors (Figure 5A).…”

Section: Resultsmentioning

confidence: 99%

“…Instead, we used HBX41108 and spautin-1 to inhibit USP7 and USP10, respectively. 24,25 Treating Caco-2bbe/NHE3 with HBX41108 or spoutin-1 decreased NHE3 protein abundance by 26.8 ± 2.2% and 14.5 ± 2.4%, respectively, whereas a 43.7 ± 1.9% reduction was achieved with both inhibitors ( Figure 5A). These results were confirmed by treating cells with a combination of one shRNA construct and an inhibitor of the other DUB.…”

Section: Usp7 and Usp10 Have An Additive Effect On Nhe3mentioning

confidence: 99%

Ubiquitin‐specific peptidase 7 (USP7) and USP10 mediate deubiquitination of human NHE3 regulating its expression and activity

Han

Yun

2020

FASEB j.

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Na + /H + exchanger NHE3 of human or primates differs from NHE3 of other animals by having a PY motif, which mediates interaction with the E3 ubiquitin (Ub) ligase Nedd4-2. Ub-conjugation of human NHE3 by Nedd4-2 regulates endocytosis of NHE3 but does not affect its cellular expression. Because Ub-conjugation is a reversible process, the aim of this study is to identify deubiquitinating enzyme (DUB) regulating the post-endosomal fate of human NHE3. Using an activity-based chemical screening, we identified ubiquitin specific protease-7 (USP7) and USP10 that bind NHE3. The roles of DUBs in regulation of NHE3 were studied by determining the effects of silencing of USP7 and USP10. Knockdown of USP7 or USP10 resulted in increased NHE3 ubiquitination and decreased NHE3 expression at the surface membrane and cellular level. The endocytic retrieval of NHE3 was promoted by depletion of USP7 or USP10, with increased association of NHE3 with Rab5a and Rab7. Inhibition of USP7 and USP10 by chemical inhibitors or knockdown had an additive effect on NHE3. In addition, NHE3 half-life was reduced accounting for decreased NHE3 protein abundance. NHE3 is inhibited by protein kinase A. Activation of PKA by forskolin decreased the binding of USP7 and USP10 to NHE3, while increasing ubiquitination of NHE3. Knockdown of USP10 had an additive effect on PKA-dependent inhibition of NHE3. These findings demonstrate that USP7 and USP10 are DUBs that regulate NHE3 ubiquitination and expression, and reveal a new mechanism of NHE3 inhibition involving DUBs. K E Y W O R D S deubiquitinating enzyme, diarrhea, Na + /H + exchange, ubiquitination How to cite this article: Han Y, Yun CC. Ubiquitinspecific peptidase 7 (USP7) and USP10 mediate deubiquitination of human NHE3 regulating its expression and activity.

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“…For instance, spautin-1 antagonizes the activity of ubiquitin specific protease 10 and ubiquitin specific protease 13 that are responsible for removing ubiquitin from the BECN1 complex. As such, spautin-1 increases the proteasome-mediated degradation of the BECN1 complex, which ultimately decreases autophagosome formation and inhibits relatively early events during autophagy ( Guo et al, 2020 ). Chloroquine is a lysosomotropic agent that accumulates in lysosomes and increases lysosomal pH to prevent lysosomal-mediated degradation.…”

Section: Discussionmentioning

confidence: 99%

Assessing methods to quantitatively validate TGFβ-dependent autophagy

Trelford

Guglielmo

2020

Biology Open

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Transforming growth factor beta (TGFβ) promotes tumorigenesis by suppressing immune surveillance and inducing epithelial to mesenchymal transition (EMT). TGFβ may augment tumorigenesis by activating autophagy, which protects cancer cells from chemotherapy and promotes invasive and anti-apoptotic properties. Here, we assess how TGFβ1 modulates autophagy related (ATG) gene expression and ATG protein levels. We also assessed microtubule-associated protein light chain 3 (LC3) lipidation, LC3 puncta formation and autophagosome-lysosome co-localization in non-small cell lung cancer (NSCLC) cell lines. These experimental approaches were validated using pharmacological autophagy inhibitors (chloroquine and spautin-1) and an autophagy activator (MG132). We found that TGFβ1, chloroquine and MG132 had little effect on ATG protein levels but increased LC3 lipidation, LC3 puncta formation and autophagosome-lysosome co-localization. Since similar outcomes were observed using chloroquine and MG132, we concluded that several techniques employed to assess TGFβ-dependent autophagy may not differentiate between the activation of autophagy versus lysosomal inhibition. Thus, NSCLC cell lines stably expressing a GFP-LC3-RFP-LC3ΔG autophagic flux probe were used to assess TGFβ-mediated autophagy. Using this approach, we observed that TGFβ, MG132 and serum starvation increased autophagic flux, whereas chloroquine and spautin-1 decreased autophagic flux. Finally, we demonstrated that ATG5 and ATG7 are critical for TGFβ-dependent autophagy in NSCLC cells. The application of this model will fuel future experiments to characterize TGFβ-dependent autophagy, which is necessary to understand the molecular processes that link, TGFβ, autophagy and tumorigenesis.

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Potent USP10/13 antagonist spautin‐1 suppresses melanoma growth via ROS‐mediated DNA damage and exhibits synergy with cisplatin

Cited by 32 publications

References 54 publications

Short linear motif candidates in the cell entry system used by SARS-CoV-2 and their potential therapeutic implications

Short linear motif candidates in the cell entry system used by SARS-CoV-2 and their potential therapeutic implications

Ubiquitin‐specific peptidase 7 (USP7) and USP10 mediate deubiquitination of human NHE3 regulating its expression and activity

Assessing methods to quantitatively validate TGFβ-dependent autophagy

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