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24 pages, 8957 KiB

Open AccessArticle

An Anti-Invasive Role for Mdmx through the RhoA GTPase under the Control of the NEDD8 Pathway

by Lara J. Bou Malhab, Susanne Schmidt, Christine Fagotto-Kaufmann, Emmanuelle Pion, Gilles Gadea, Pierre Roux, Francois Fagotto, Anne Debant and Dimitris P. Xirodimas

Cells 2024, 13(19), 1625; https://doi.org/10.3390/cells13191625 - 28 Sep 2024

Viewed by 548

Abstract

Mdmx (Mdm4) is established as an oncogene mainly through repression of the p53 tumour suppressor. On the other hand, anti-oncogenic functions for Mdmx have also been proposed, but the underlying regulatory pathways remain unknown. Investigations into the effect of inhibitors for the NEDD8 [...] Read more.

Mdmx (Mdm4) is established as an oncogene mainly through repression of the p53 tumour suppressor. On the other hand, anti-oncogenic functions for Mdmx have also been proposed, but the underlying regulatory pathways remain unknown. Investigations into the effect of inhibitors for the NEDD8 pathway in p53 activation, human cell morphology, and in cell motility during gastrulation in Xenopus embryos revealed an anti-invasive function of Mdmx. Through stabilisation and activation of the RhoA GTPase, Mdmx is required for the anti-invasive effects of NEDDylation inhibitors. Mechanistically, through its Zn finger domain, Mdmx preferentially interacts with the inactive GDP-form of RhoA. This protects RhoA from degradation and allows for RhoA targeting to the plasma membrane for its subsequent activation. The effect is transient, as prolonged NEDDylation inhibition targets Mdmx for degradation, which subsequently leads to RhoA destabilisation. Surprisingly, Mdmx degradation requires non-NEDDylated (inactive) Culin4A and the Mdm2 E3-ligase. This study reveals that Mdmx can control cell invasion through RhoA stabilisation/activation, which is potentially linked to the reported anti-oncogenic functions of Mdmx. As inhibitors of the NEDD8 pathway are in clinical trials, the status of Mdmx may be a critical determinant for the anti-tumour effects of these inhibitors. Full article

(This article belongs to the Special Issue Roles and Mechanisms of Ubiquitin Ligases (E3) and Deubiquitinases (DUBs))

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17 pages, 18244 KiB

Open AccessArticle

Structural Dynamics Analysis of USP14 Activation by AKT-Mediated Phosphorylation

by Raju Dash, Non-Nuoc Tran, Sung Bae Lee and Byung-Hoon Lee

Cells 2024, 13(11), 955; https://doi.org/10.3390/cells13110955 - 31 May 2024

Viewed by 794

Abstract

Ubiquitin-specific protease 14 (USP14), one of the three major proteasome-associated deubiquitinating enzymes (DUBs), is known to be activated by the AKT-mediated phosphorylation at Ser432. Thereby, AKT can regulate global protein degradation by controlling the ubiquitin-proteasome system (UPS). However, the exact molecular mechanism of [...] Read more.

Ubiquitin-specific protease 14 (USP14), one of the three major proteasome-associated deubiquitinating enzymes (DUBs), is known to be activated by the AKT-mediated phosphorylation at Ser432. Thereby, AKT can regulate global protein degradation by controlling the ubiquitin-proteasome system (UPS). However, the exact molecular mechanism of USP14 activation by AKT phosphorylation at the atomic level remains unknown. By performing the molecular dynamics (MD) simulation of the USP14 catalytic domain at three different states (inactive, active, and USP14-ubiquitin complex), we characterized the change in structural dynamics by phosphorylation. We observed that the Ser432 phosphorylation induced substantial conformational changes of USP14 in the blocking loop (BL) region to fold it from an open loop into a β-sheet, which is critical for USP14 activation. Furthermore, phosphorylation also increased the frequency of critical hydrogen bonding and salt bridge interactions between USP14 and ubiquitin, which is essential for DUB activity. Structural dynamics insights from this study pinpoint the important local conformational landscape of USP14 by the phosphorylation event, which would be critical for understanding USP14-mediated proteasome regulation and designing future therapeutics. Full article

(This article belongs to the Special Issue Roles and Mechanisms of Ubiquitin Ligases (E3) and Deubiquitinases (DUBs))

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19 pages, 3914 KiB

Open AccessArticle

The E3 Ubiquitin Protein Ligase LINCR Amplifies the TLR-Mediated Signals through Direct Degradation of MKP1

by Takumi Yokosawa, Sayoko Miyagawa, Wakana Suzuki, Yuki Nada, Yusuke Hirata, Takuya Noguchi and Atsushi Matsuzawa

Cells 2024, 13(8), 687; https://doi.org/10.3390/cells13080687 - 15 Apr 2024

Viewed by 1178

Abstract

Toll-like receptors (TLRs) induce innate immune responses through activation of intracellular signaling pathways, such as MAP kinase and NF-κB signaling pathways, and play an important role in host defense against bacterial or viral infections. Meanwhile, excessive activation of TLR signaling leads to a [...] Read more.

Toll-like receptors (TLRs) induce innate immune responses through activation of intracellular signaling pathways, such as MAP kinase and NF-κB signaling pathways, and play an important role in host defense against bacterial or viral infections. Meanwhile, excessive activation of TLR signaling leads to a variety of inflammatory disorders, including autoimmune diseases. TLR signaling is therefore strictly controlled to balance optimal immune response and inflammation. However, its balancing mechanisms are not fully understood. In this study, we identified the E3 ubiquitin ligase LINCR/ NEURL3 as a critical regulator of TLR signaling. In LINCR-deficient cells, the sustained activation of JNK and p38 MAPKs induced by the agonists for TLR3, TLR4, and TLR5, was clearly attenuated. Consistent with these observations, TLR-induced production of a series of inflammatory cytokines was significantly attenuated, suggesting that LINCR positively regulates innate immune responses by promoting the activation of JNK and p38. Interestingly, our further mechanistic study identified MAPK phosphatase-1 (MKP1), a negative regulator of MAP kinases, as a ubiquitination target of LINCR. Thus, our results demonstrate that TLRs fine-tune the activation of MAP kinase pathways by balancing LINCR (the positive regulator) and MKP1 (the negative regulator), which may contribute to the induction of optimal immune responses. Full article

(This article belongs to the Special Issue Roles and Mechanisms of Ubiquitin Ligases (E3) and Deubiquitinases (DUBs))

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19 pages, 3690 KiB

Open AccessArticle

UBE3C Facilitates the ER-Associated and Peripheral Degradation of Misfolded CFTR

by Yuka Kamada, Hazuki Tateishi, Uta Nakayamada, Daichi Hinata, Ayuka Iwasaki, Jingxin Zhu, Ryosuke Fukuda and Tsukasa Okiyoneda

Cells 2023, 12(23), 2741; https://doi.org/10.3390/cells12232741 - 30 Nov 2023

Cited by 3 | Viewed by 1620

Abstract

The ubiquitin E3 ligase UBE3C promotes the proteasomal degradation of cytosolic proteins and endoplasmic reticulum (ER) membrane proteins. UBE3C is proposed to function downstream of the RNF185/MBRL ER-associated degradation (ERAD) branch, contributing to the ERAD of select membrane proteins. Here, we report that [...] Read more.

The ubiquitin E3 ligase UBE3C promotes the proteasomal degradation of cytosolic proteins and endoplasmic reticulum (ER) membrane proteins. UBE3C is proposed to function downstream of the RNF185/MBRL ER-associated degradation (ERAD) branch, contributing to the ERAD of select membrane proteins. Here, we report that UBE3C facilitates the ERAD of misfolded CFTR, even in the absence of both RNF185 and its functional ortholog RNF5 (RNF5/185). Unlike RNF5/185, UBE3C had a limited impact on the ubiquitination of misfolded CFTR. UBE3C knockdown (KD) resulted in an additional increase in the functional ∆F508-CFTR channels on the plasma membrane when combined with the RNF5/185 ablation, particularly in the presence of clinically used CFTR modulators. Interestingly, although UBE3C KD failed to attenuate the ERAD of insig-1, it reduced the ERAD of misfolded ∆Y490-ABCB1 and increased cell surface expression. UBE3C KD also stabilized the mature form of ∆F508-CFTR and increased the cell surface level of T70-CFTR, a class VI CFTR mutant. These results suggest that UBE3C plays a vital role in the ERAD of misfolded CFTR and ABCB1, even within the RNF5/185-independent ERAD pathway, and it may also be involved in maintaining the peripheral quality control of CFTR. Full article

(This article belongs to the Special Issue Roles and Mechanisms of Ubiquitin Ligases (E3) and Deubiquitinases (DUBs))

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Review

Jump to: Research

30 pages, 10423 KiB

Open AccessReview

Harmony of Protein Tags and Chimeric Molecules Empowers Targeted Protein Ubiquitination and Beyond

by Aggie Lawer, Luke Schulz, Renata Sawyer and Xuyu Liu

Cells 2024, 13(5), 426; https://doi.org/10.3390/cells13050426 - 28 Feb 2024

Viewed by 2264

Abstract

Post-translational modifications (PTMs) are crucial mechanisms that underlie the intricacies of biological systems and disease mechanisms. This review focuses on the latest advancements in the design of heterobifunctional small molecules that hijack PTM machineries for target-specific modifications in living systems. A key innovation [...] Read more.

Post-translational modifications (PTMs) are crucial mechanisms that underlie the intricacies of biological systems and disease mechanisms. This review focuses on the latest advancements in the design of heterobifunctional small molecules that hijack PTM machineries for target-specific modifications in living systems. A key innovation in this field is the development of proteolysis-targeting chimeras (PROTACs), which promote the ubiquitination of target proteins for proteasomal degradation. The past decade has seen several adaptations of the PROTAC concept to facilitate targeted (de)phosphorylation and acetylation. Protein fusion tags have been particularly vital in these proof-of-concept studies, aiding in the investigation of the functional roles of post-translationally modified proteins linked to diseases. This overview delves into protein-tagging strategies that enable the targeted modulation of ubiquitination, phosphorylation, and acetylation, emphasizing the synergies and challenges of integrating heterobifunctional molecules with protein tags in PTM research. Despite significant progress, many PTMs remain to be explored, and protein tag-assisted PTM-inducing chimeras will continue to play an important role in understanding the fundamental roles of protein PTMs and in exploring the therapeutic potential of manipulating protein modifications, particularly for targets not yet addressed by existing drugs. Full article

(This article belongs to the Special Issue Roles and Mechanisms of Ubiquitin Ligases (E3) and Deubiquitinases (DUBs))

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10 pages, 785 KiB

Open AccessReview

E3 Ligases Regulate Organelle Inheritance in Yeast

by Keisuke Obara, Kohei Nishimura and Takumi Kamura

Cells 2024, 13(4), 292; https://doi.org/10.3390/cells13040292 - 6 Feb 2024

Cited by 1 | Viewed by 1410

Abstract

Saccharomyces cerevisiae proliferates by budding, which includes the formation of a cytoplasmic protrusion called the ‘bud’, into which DNA, RNA, proteins, organelles, and other materials are transported. The transport of organelles into the growing bud must be strictly regulated for the proper inheritance [...] Read more.

Saccharomyces cerevisiae proliferates by budding, which includes the formation of a cytoplasmic protrusion called the ‘bud’, into which DNA, RNA, proteins, organelles, and other materials are transported. The transport of organelles into the growing bud must be strictly regulated for the proper inheritance of organelles by daughter cells. In yeast, the RING-type E3 ubiquitin ligases, Dma1 and Dma2, are involved in the proper inheritance of mitochondria, vacuoles, and presumably peroxisomes. These organelles are transported along actin filaments toward the tip of the growing bud by the myosin motor protein, Myo2. During organelle transport, organelle-specific adaptor proteins, namely Mmr1, Vac17, and Inp2 for mitochondria, vacuoles, and peroxisomes, respectively, bridge the organelles and myosin. After reaching the bud, the adaptor proteins are ubiquitinated by the E3 ubiquitin ligases and degraded by the proteasome. Targeted degradation of the adaptor proteins is necessary to unload vacuoles, mitochondria, and peroxisomes from the actin–myosin machinery. Impairment of the ubiquitination of adaptor proteins results in the failure of organelle release from myosin, which, in turn, leads to abnormal dynamics, morphology, and function of the inherited organelles, indicating the significance of proper organelle unloading from myosin. Herein, we summarize the role and regulation of E3 ubiquitin ligases during organelle inheritance in yeast. Full article

(This article belongs to the Special Issue Roles and Mechanisms of Ubiquitin Ligases (E3) and Deubiquitinases (DUBs))

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12 pages, 748 KiB

Open AccessReview

Targeted Protein Degradation Systems: Controlling Protein Stability Using E3 Ubiquitin Ligases in Eukaryotic Species

by Yoshitaka Ogawa, Taisei P. Ueda, Keisuke Obara, Kohei Nishimura and Takumi Kamura

Cells 2024, 13(2), 175; https://doi.org/10.3390/cells13020175 - 17 Jan 2024

Cited by 1 | Viewed by 2370

Abstract

This review explores various methods for modulating protein stability to achieve target protein degradation, which is a crucial aspect in the study of biological processes and drug design. Thirty years have passed since the introduction of heat-inducible degron cells utilizing the N-end rule, [...] Read more.

This review explores various methods for modulating protein stability to achieve target protein degradation, which is a crucial aspect in the study of biological processes and drug design. Thirty years have passed since the introduction of heat-inducible degron cells utilizing the N-end rule, and methods for controlling protein stability using the ubiquitin–proteasome system have moved from academia to industry. This review covers protein stability control methods, from the early days to recent advancements, and discusses the evolution of techniques in this field. This review also addresses the challenges and future directions of protein stability control techniques by tracing their development from the inception of protein stability control methods to the present day. Full article

(This article belongs to the Special Issue Roles and Mechanisms of Ubiquitin Ligases (E3) and Deubiquitinases (DUBs))

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19 pages, 2871 KiB

Open AccessReview

Role of Ubiquitination and Epigenetics in the Regulation of AhR Signaling in Carcinogenesis and Metastasis: “Albatross around the Neck” or “Blessing in Disguise”

by Ammad Ahmad Farooqi, Venera Rakhmetova, Gulnara Kapanova, Gulnur Tanbayeva, Akmaral Mussakhanova, Akmaral Abdykulova and Alma-Gul Ryskulova

Cells 2023, 12(19), 2382; https://doi.org/10.3390/cells12192382 - 29 Sep 2023

Cited by 2 | Viewed by 2062

Abstract

The molecular mechanisms and signal transduction cascades evoked by the activation of aryl hydrocarbon receptor (AhR) are becoming increasingly understandable. AhR is a ligand-activated transcriptional factor that integrates environmental, dietary and metabolic cues for the pleiotropic regulation of a wide variety of mechanisms. [...] Read more.

The molecular mechanisms and signal transduction cascades evoked by the activation of aryl hydrocarbon receptor (AhR) are becoming increasingly understandable. AhR is a ligand-activated transcriptional factor that integrates environmental, dietary and metabolic cues for the pleiotropic regulation of a wide variety of mechanisms. AhR mediates transcriptional programming in a ligand-specific, context-specific and cell-type-specific manner. Pioneering cutting-edge research works have provided fascinating new insights into the mechanistic role of AhR-driven downstream signaling in a wide variety of cancers. AhR ligands derived from food, environmental contaminants and intestinal microbiota strategically activated AhR signaling and regulated multiple stages of cancer. Although AhR has classically been viewed and characterized as a ligand-regulated transcriptional factor, its role as a ubiquitin ligase is fascinating. Accordingly, recent evidence has paradigmatically shifted our understanding and urged researchers to drill down deep into these novel and clinically valuable facets of AhR biology. Our rapidly increasing realization related to AhR-mediated regulation of the ubiquitination and proteasomal degradation of different proteins has started to scratch the surface of intriguing mechanisms. Furthermore, AhR and epigenome dynamics have shown previously unprecedented complexity during multiple stages of cancer progression. AhR not only transcriptionally regulated epigenetic-associated molecules, but also worked with epigenetic-modifying enzymes during cancer progression. In this review, we have summarized the findings obtained not only from cell-culture studies, but also from animal models. Different clinical trials are currently being conducted using AhR inhibitors and PD-1 inhibitors (Pembrolizumab and nivolumab), which confirm the linchpin role of AhR-related mechanistic details in cancer progression. Therefore, further studies are required to develop a better comprehension of the many-sided and “diametrically opposed” roles of AhR in the regulation of carcinogenesis and metastatic spread of cancer cells to the secondary organs. Full article

(This article belongs to the Special Issue Roles and Mechanisms of Ubiquitin Ligases (E3) and Deubiquitinases (DUBs))

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