Synthetic Lethal Interaction between the ESCRT Paralog Enzymes VPS4A and VPS4B in Cancers Harboring Loss of Chromosome 18q or 16q

Neggers, Jasper E.; Paolella, Brenton R.; Asfaw, Adhana; Rothberg, Michael; Skipper, Thomas A.; Yang, Annan; Kalekar, Radha L.; Krill-Burger, John M.; Dharia, Neekesh V.; Kugener, Guillaume; Kalfon, Jérémie; Chen, Yuan; Dumont, Nancy; González, Alfredo Bullard; Abdusamad, Mai; Li, Yvonne Y.; Wu, Westley W.; Durbin, Adam D.; Wolpin, Brian M.; Piccioni, Federica; Root, David E.; Boehm, Jesse S.; Cherniack, Andrew D.; Tsherniak, Aviad; Hong, Andrew L.; Hahn, William C.; Stegmaier, Kimberly; Golub, Todd R.; Vázquez, Francisca; Aguirre, Andrew J.

doi:10.1016/j.celrep.2021.109367

Cited by 5 publications

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“…We next performed similar experiments testing a total of 16 unique POIs, spanning different protein classes including ten cytoplasmic proteins (Green Fluorescent Protein (GFP), NanoLuciferase (NanoLuc), Red-shifted Firefly Luciferase (RFluc 18 ), Protein arginine methyltransferase 2 (PRMT5 19 ), WD Repeat and SOCS Box-Containing 2 (WSB2), induced myeloid leukemia cell differentiation protein (MCL1 20 ), Soc-2 Suppressor Of Clear Homolog (SHOC2 21,22 ), Methionine Adenosyltransferase 2 A (MAT2A 23 ), Vacuolar protein sortingassociated protein 4 (VPS4A 24 ), and Protein Activator of interferoninduced Protein Kinase EIF2AK2 (PRKRA 25 ); three nuclear proteins (Stromal Antigen 1 and 2 (STAG1 and STAG2 26 ), DNA (cytosine-5)methyltransferase 1 (DNMT3A 27 ); two multi-pass plasma membrane proteins (Xenotropic and Polytropic Receptor 1 (XPR1 28,29 ), and Kinase D interacting protein of 220 kDa (KIDINS220 30,31 ); and one single-pass transmembrane receptor tyrosine kinase (Fms-Related receptor tyrosine kinase 3 with the internal tandem duplication, FLT3-ITD 27 ).…”

Section: Cdt Performance Across 16 Unique Poismentioning

confidence: 99%

Systematic profiling of conditional degron tag technologies for target validation studies

et al. 2022

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Conditional degron tags (CDTs) are a powerful tool for target validation that combines the kinetics and reversible action of pharmacological agents with the generalizability of genetic manipulation. However, successful design of a CDT fusion protein often requires a prolonged, ad hoc cycle of construct design, failure, and re-design. To address this limitation, we report here a system to rapidly compare the activity of five unique CDTs: AID/AID2, IKZF3d, dTAG, HaloTag, and SMASh. We demonstrate the utility of this system against 16 unique protein targets. We find that expression and degradation are highly dependent on the specific CDT, the construct design, and the target. None of the CDTs leads to efficient expression and/or degradation across all targets; however, our systematic approach enables the identification of at least one optimal CDT fusion for each target. To enable the adoption of CDT strategies more broadly, we have made these reagents, and a detailed protocol, available as a community resource.

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Section: Cdt Performance Across 16 Unique Poismentioning

confidence: 99%

Systematic profiling of conditional degron tag technologies for target validation studies

et al. 2022

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“…Vps4b, which is another key ESCRT family protein, is structurally and functionally similar to Vps4a, being involved in the depolymerization of ESCRT III. In tumor cells, knockout of Vps4b led to an increased expression of Vps4a to compensate for these functions [ 32 , 33 ]. The RNA sequencing results from this study also showed that Vps4b expression was increased in Vps4a knockout mice ( Figure 3 C).…”

Section: Discussionmentioning

confidence: 99%

“…The fact that other mice did not die shows what the process of sealing autophagosomes can accomplish. There is synthetic lethality between Vps4a and Vps4b [ 32 , 33 ]. Knockdown of Vps4a alone was not sufficient to cause a complete loss of membrane remodeling mediated by ESCRT.…”

Section: Discussionmentioning

confidence: 99%

Vps4a Regulates Autophagic Flux to Prevent Hypertrophic Cardiomyopathy

Huang

Zhang

Wang

et al. 2023

IJMS

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Autophagy has stabilizing functions for cardiomyocytes. Recent studies indicate that an impairment in the autophagy pathway can seriously affect morphology and function, potentially leading to heart failure. However, the role and the underlying mechanism of the endosomal sorting complex required for transport (ESCRT) family protein, in particular the AAA-ATPase vacuolar protein sorting 4a (Vps4a), in regulating myocardial autophagy remains unclear. In the present study, cardiomyocyte-specific Vps4a knockout mice were generated by crossing Vps4aflox/flox (Vps4afl/fl) with Myh6-cre transgenic mice. As a result, we observed a partially dilated left ventricular (LV) chamber, a significant increase in heart weight to body weight ratio (HW/BW), and heart weight to tibial length ratio (HW/TL), hypertrophic cardiomyopathy and early lethality starting at 3 months of age. Hematoxylin-eosin (HE), immunofluorescence assay (IFA), and Western blot (WB) revealed autophagosome accumulation in cardiomyocytes. A transcriptome-based analysis and autophagic flux tracking by AAV-RFP-GFP-LC3 showed that the autophagic flux was blocked in Vps4a knockout cardiomyocytes. In addition, we provided in vitro evidence demonstrating that Vps4a and LC3 were partially co-localized in cardiomyocytes, and the knockdown of Vps4a led to the accumulation of autophagosomes in cardiomyocytes. Similarly, the transfection of cardiomyocytes with adenovirus (Adv) mCherry-GFP-LC3 further indicated that the autophagic flux was blocked in cells with deficient levels of Vps4a. Finally, an electron microscope (EM) showed that the compromised sealing of autophagosome blocked the autophagic flux in Vps4a-depleted cardiomyocytes. These findings revealed that Vps4a contributed to the sealing of autophagosomes in cardiomyocytes. Therefore, we demonstrated that Vps4a deletion could block the autophagic flux, leading to the accumulation of degradation substances and compromised cardiac function. Overall, this study provides insights into a new theoretical basis for which autophagy may represent a therapeutic target for cardiovascular diseases.

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“…Our observation that CHMP7 interacts exclusively with the MIT domain of the AMSH deubiquitinase supports the possibility that these events may be dynamically regulated by ubiquitin-dependent processes, as has been recently reported(Wallis et al, 2021). More generally, our quantitative definition of the ESCRT-III-MIT interactome should provide a basis for probing how disruption of ESCRT-III and MIT cofactor activities can contribute to disease states such as hereditary spastic paraplegia(Ciccarelli et al, 2003), or can be used therapeutically, for example in anti-cancer strategies based on VPS4 synthetic lethality(Neggers et al, 2020, Szymanska et al, 2020).…”

Section: Discussionmentioning

confidence: 99%

Comprehensive analysis of the human ESCRT-III-MIT domain interactome reveals new cofactors for cytokinetic abscission

Wenzel

Mackay

Skalicky

et al. 2022

Preprint

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The 12 related human ESCRT-III proteins form filaments that constrict membranes and mediate fission, including during cytokinetic abscission. The C-terminal tails of polymerized ESCRT-III subunits also bind proteins that contain Microtubule-Interacting and Trafficking (MIT) domains. MIT domains can interact with ESCRT-III tails in many different ways to create a complex binding code that recruits essential cofactors to sites of ESCRT activity. Here, we have comprehensively and quantitatively mapped the interactions between all known ESCRT-III tails and 19 pure recombinant human MIT domains. We measured 228 pairwise interactions, quantified 58 positive interactions, and discovered 16 previously unreported interactions. We also report the crystal structure of the SPASTIN MIT domain in complex with the IST1 C-terminal tail. Three MIT enzymes were studied in detail and shown to: 1) localize to cytokinetic midbody membrane bridges through interactions with their specific ESCRT-III binding partners (SPASTIN-IST1, KATNA1-CHMP3, and CAPN7-IST1), 2) function in abscission (SPASTIN, KATNA1, and CAPN7), and 3) function in the 'NoCut' abscission checkpoint (SPASTIN and CAPN7). Our studies define the human MIT-ESCRT-III interactome, identify new factors and activities required for cytokinetic abscission and its regulation, and provide a platform for analyzing ESCRT-III and MIT cofactor interactions in all ESCRT-mediated processes.

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Synthetic Lethal Interaction between the ESCRT Paralog Enzymes VPS4A and VPS4B in Cancers Harboring Loss of Chromosome 18q or 16q

Abstract: In the originally published version of this paper, the CHMP4B sgRNA sequence listed in the STAR Methods section was incorrect. The incorrect sequence was 5 0 -TCGATGGCACAAGCCATGAA, which is an sgRNA designed to target CHMP2A.

Cited by 5 publications

References 0 publications

Systematic profiling of conditional degron tag technologies for target validation studies

Systematic profiling of conditional degron tag technologies for target validation studies

Vps4a Regulates Autophagic Flux to Prevent Hypertrophic Cardiomyopathy

Comprehensive analysis of the human ESCRT-III-MIT domain interactome reveals new cofactors for cytokinetic abscission

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