Assessing the mechanism of fast‐cycling cancer‐associated mutations of Rac1 small Rho GTPase

Protein Science, Journal Year: 2024, Volume and Issue: 33(4)

Published: March 19, 2024

Rho-GTPases proteins function as molecular switches alternating from an active to inactive state upon Guanosine triphosphate (GTP) binding and hydrolysis diphosphate (GDP). Among them, Rac subfamily regulates cell dynamics, being overexpressed in distinct cancer types. Notably, these are object of frequent cancer-associated mutations at Pro29 (P29S, P29L, P29Q). To assess the impact on Rac1 structure function, we performed extensive all-atom dynamics simulations wild-type (wt) oncogenic isoforms this protein GDP- GTP-bound states. Our results unprecedentedly elucidate that P29Q/S-induced structural dynamical perturbations core domain weaken catalytic site Mg

Language: Английский

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Molecular dynamics simulations for the structure-based drug design: targeting small-GTPases proteins

Expert Opinion on Drug Discovery, Journal Year: 2024, Volume and Issue: 19(10), P. 1259 - 1279

Published: Aug. 6, 2024

Molecular Dynamics (MD) simulations can support mechanism-based drug design. Indeed, MD by capturing biomolecule motions at finite temperatures reveal hidden binding sites, accurately predict drug-binding poses, and estimate the thermodynamics kinetics, crucial information for discovery campaigns. Small-Guanosine Triphosphate Phosphohydrolases (GTPases) regulate a cascade of signaling events, that affect most cellular processes. Their deregulation is linked to several diseases, making them appealing targets. The broad roles small-GTPases in processes recent approval covalent KRas inhibitor as an anticancer agent renewed interest targeting small-GTPase with small molecules.

Language: Английский

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New insights into the classification of the RAC1 P29S hotspot mutation in melanoma as an oncogene

Research Square (Research Square), Journal Year: 2025, Volume and Issue: unknown

Published: March 27, 2025

The RAC1^P29S hotspot mutation, prevalent in melanoma, drives tumorigenesis by enhancing molecular interactions and hyperactivating key signaling pathways, making it a compelling target for cancer therapy. This study provides comprehensive biochemical characterization of RAC1^P29S compared to wild-type RAC1 mutations T17N F28L. P29S mutation significantly impairs nucleotide binding guanosine triphosphate (GTP) diphosphate, accelerating intrinsic exchange. While minimally affecting regulation dissociation inhibitor 1, exhibits reduced activation via diffuse B-cell lymphoma family guanine exchange factors but retains effective dedicator cytokinesis 2. Critically, the severely GTPase-activating protein-stimulated GTP hydrolysis, most likely contributing hyperactivation prolonging its GTP-bound form. displays stronger affinity IQ motif-containing protein 1 than p21-activated kinase highlighting role former scaffolding RAC1^P29S-driven signaling. In serum-starved cells, predominantly adopts an active state. overexpression activates cancer-associated including extracellular signal-regulated p38 mitogen-activated kinase, reinforcing as oncogenic driver melanoma. These insights suggest potential therapeutic targets melanoma treatment, regulators modulators.

Language: Английский

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Quantifying small GTPase activation status using a novel fluorescence HPLC-based assay

Journal of Biological Chemistry, Journal Year: 2025, Volume and Issue: unknown, P. 108545 - 108545

Published: April 1, 2025

Small GTPases play crucial roles in cellular signaling pathways, with their activation states tightly regulated between GDP-bound inactive and GTP-bound active forms. Dysregulation of these nucleotide-binding states, such as oncogenic RAS, is implicated diseases like cancer. Accurately quantifying cells thus for deciphering functional regulatory mechanisms. However, current methods do not fully meet the necessary sensitivity versatility, limiting effectiveness small GTPase analysis. Here, we present a highly sensitive HPLC-based assay fluorescence detection (Fluor-HPLC), enabling precise quantification guanine GTPases. Applying this technique, successfully quantified at endogenous expression levels. We demonstrated utility Fluor-HPLC by elucidating RHEB HRAS response to extracellular stimuli. Furthermore, integration syngeneic mouse models provided insights into KRAS dynamics tumor tissues evaluated targeted therapeutics. Overall, versatile method paves way investigating mechanisms various GTPases, potentially accelerating our understanding processes disease pathogenesis.

Language: Английский

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Assessing the Mechanism of Rac1b: An All-Atom Simulation Study of the Alternative Spliced Variant of Rac1 Small Rho GTPase

Journal of Chemical Information and Modeling, Journal Year: 2024, Volume and Issue: unknown

Published: Dec. 5, 2024

The Rho GTPase family plays a key role in cell migration, cytoskeletal dynamics, and intracellular signaling. Rac1 its splice variant Rac1b, characterized by the insertion of an Extraloop, are frequently associated with cancer. These small GTPases switch between active GTP-bound state inactive GDP-bound state, process that is regulated specific protein modulators. Among them, Guanine nucleotide exchange factor (GEF) DOCK5 specifically targets GTPases, promoting their activation facilitating GDP for GTP. In this study, we performed cumulative 10-μs-long all-atom molecular dynamics simulations isolation complex ELMO1, to investigate impact Rac1b Extraloop. Our findings reveal Extraloop decreases residence time as compared Rac1, mimicking effect accelerated GDP/GTP induced DOCK5. Furthermore, both ELMO1 stabilize GTPase/DOCK5 complex, contributing facilitate dissociation. This shifts balance GPT- toward sending prooncogenic signal. Besides broadening our understanding biological functions study provides information exploit previously unexplored therapeutic niche counter Rac1b-associated

Language: Английский

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ZIP13 regulates lipid metabolism by changing intracellular iron and zinc balance

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: July 5, 2024

Abstract Metabolic diseases are caused by a prolonged energy imbalance, and adipose tissue is known to be the main contributor. We previously reported that ZIP13, an Slc39a transporter whose deficiency causes Ehlers-Danlos syndrome spondylocheirodysplastic type 3 associated with lipoatrophy, inhibits adipocyte browning pathway modulating intracellular zinc status. The precise mechanisms of how ZIP13 regulates homeostasis remain unclear therefore, we investigated role in mature adipocytes using adipocyte-specific Zip13 -deficient mice. herein demonstrate these mice show accelerated lipolysis reduced respiratory exchange ratio. In addition, abundance iron balance were altered during differentiation normal adipocytes, whereas distribution was substantially affected which downregulated PDE activity enhanced β-adrenergic receptor signaling pathways. Importantly, confirmed could transport both iron, Xenopus oocyte system silico structural dynamics simulations, defect perturbs proper lipolysis. Together, results illustrate acts as key regulator for via use metals, ZIP13-iron axis plays important regulation lipid metabolism.

Language: Английский

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Structural dynamics of Rho GTPases

Journal of Molecular Biology, Journal Year: 2024, Volume and Issue: unknown, P. 168919 - 168919

Published: Dec. 1, 2024

Language: Английский

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