A genome‐wide <scp>CRISPR</scp> screen identifies the <scp>CCT</scp> chaperonin as a critical regulator of vesicle trafficking

Chen, Yongtian; Kang, Jing; Zhen, Ru; Zhang, Jintao; Chen, Caiyong

doi:10.1096/fj.202201580r

Cited by 4 publications

(2 citation statements)

References 56 publications

(131 reference statements)

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“…We detected multiple components of the TRiC molecular chaperone complex (CCT8, TCP1, CCT6A, CCT3 and CCT4), that was also enriched as GO term (Supplementary Fig. 10d; Supplementary Data 8) and, in fact, CCT4 was recently implicated as a crucial vesicular trafficking regulator 56 . We validated CCT8, as well as TP53BP2, as NEDD4 Ub substrates by WB (Fig.…”

Section: Bioe3 Identifies Nedd4 Substratesmentioning

confidence: 99%

BioE3 enables the identification ofbona fidetargets of E3 ligases

Barroso-Gomila,

Merino-Cacho,

Muratore

et al. 2023

Preprint

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The post-translational modification of proteins by ubiquitination is a highly regulated process that involves a dynamic, three-step enzymatic cascade, where more than 600 E3 ligases play a critical role in recognizing specific substrates for modification. Separating bona fide targets of E3s from E3-interacting proteins remains a major challenge in the field. In this study, we present BioE3, a novel approach for identifying substrates of ubiquitin-like (UbL) E3 ligases of interest. Using BirA-E3 ligase fusion proteins and bioUbLs, the method facilitates site-specific biotinylation of UbL-modified substrates of particular E3s for proteomic identification. We demonstrate that the BioE3 system can identify both known and novel targets of two RING-type ubiquitin E3 ligases: RNF4, known to be involved in DNA damage 36 response and the regulation of PML nuclear bodies, and MIB1, implicated in endocytosis, autophagy, and centrosomal protein homeostasis. We further show the versatility of BioE3 by identifying targets of an organelle-specific E3 (MARCH5) and a relatively uncharacterized E3 (RNF214). Furthermore, we show that BioE3 works with HECT-type E3 ligases and identify novel targets of NEDD4 involved in vesicular trafficking. BioE3 is a powerful tool that enables identification of bona fide substrates of UbL E3 ligases and how they change with chemical perturbations. BioE3 may also be applicable for UbLs beyond Ub and SUMO, as well as other E3 ligase classes. The resulting knowledge can shed light on the regulation of cellular processes by the complex UbL network and provide information useful for strategies such as targeted protein degradation (TPD), advancing our understanding of fundamental biological mechanisms and their applications.

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Section: Bioe3 Identifies Nedd4 Substratesmentioning

confidence: 99%

BioE3 enables the identification ofbona fidetargets of E3 ligases

Barroso-Gomila,

Merino-Cacho,

Muratore

et al. 2023

Preprint

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“…They consist of a spherical proteolipid bilayer membrane, and contain specific proteins, RNA molecules and DNA molecules 144 . Recent studies have demonstrated that CCT4 is a key regulator of vesicle trafficking, 145 and the levels of CCT subunits were found to be upregulated in various cancer exosomes 146,147 . Additionally, the levels of six of the eight CCT subunits (CCT1, CCT2, CCT3, CCT5, CCT6A and CCT7) were found to be significantly increased in glioblastoma cell‐derived extracellular vesicles, and CCT6A in glioblastoma tissue showed a potential link with EGFR and was identified as a potential prognostic biomarker 146 .…”

Section: Cct Subunits In Cancersmentioning

confidence: 99%

Revisiting the chaperonin T‐complex protein‐1 ring complex in human health and disease: A proteostasis modulator and beyond

Zeng,

Han,

Pan

et al. 2024

Clinical & Translational Med

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BackgroundDisrupted protein homeostasis (proteostasis) has been demonstrated to facilitate the progression of various diseases. The cytosolic T‐complex protein‐1 ring complex (TRiC/CCT) was discovered to be a critical player in orchestrating proteostasis by folding eukaryotic proteins, guiding intracellular localisation and suppressing protein aggregation. Intensive investigations of TRiC/CCT in different fields have improved the understanding of its role and molecular mechanism in multiple physiological and pathological processes.Main bodyIn this review, we embark on a journey through the dynamic protein folding cycle of TRiC/CCT, unraveling the intricate mechanisms of its substrate selection, recognition, and intriguing folding and assembly processes. In addition to discussing the critical role of TRiC/CCT in maintaining proteostasis, we detail its involvement in cell cycle regulation, apoptosis, autophagy, metabolic control, adaptive immunity and signal transduction processes. Furthermore, we meticulously catalogue a compendium of TRiC‐associated diseases, such as neuropathies, cardiovascular diseases and various malignancies. Specifically, we report the roles and molecular mechanisms of TRiC/CCT in regulating cancer formation and progression. Finally, we discuss unresolved issues in TRiC/CCT research, highlighting the efforts required for translation to clinical applications, such as diagnosis and treatment.ConclusionThis review aims to provide a comprehensive view of TRiC/CCT for researchers to inspire further investigations and explorations of potential translational possibilities.

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BioE3 identifies specific substrates of ubiquitin E3 ligases

Barroso-Gomila,

Merino-Cacho,

Muratore

et al. 2023

Nat Commun

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Hundreds of E3 ligases play a critical role in recognizing specific substrates for modification by ubiquitin (Ub). Separating genuine targets of E3s from E3-interactors remains a challenge. We present BioE3, a powerful approach for matching substrates to Ub E3 ligases of interest. Using BirA-E3 ligase fusions and bioUb, site-specific biotinylation of Ub-modified substrates of particular E3s facilitates proteomic identification. We show that BioE3 identifies both known and new targets of two RING-type E3 ligases: RNF4 (DNA damage response, PML bodies), and MIB1 (endocytosis, autophagy, centrosome dynamics). Versatile BioE3 identifies targets of an organelle-specific E3 (MARCH5) and a relatively uncharacterized E3 (RNF214). Furthermore, BioE3 works with NEDD4, a HECT-type E3, identifying new targets linked to vesicular trafficking. BioE3 detects altered specificity in response to chemicals, opening avenues for targeted protein degradation, and may be applicable for other Ub-likes (UbLs, e.g., SUMO) and E3 types. BioE3 applications shed light on cellular regulation by the complex UbL network.

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A genome‐wide CRISPR screen identifies the CCT chaperonin as a critical regulator of vesicle trafficking

Cited by 4 publications

References 56 publications

BioE3 enables the identification ofbona fidetargets of E3 ligases

BioE3 enables the identification ofbona fidetargets of E3 ligases

Revisiting the chaperonin T‐complex protein‐1 ring complex in human health and disease: A proteostasis modulator and beyond

BioE3 identifies specific substrates of ubiquitin E3 ligases

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