Chemoproteomic-enabled characterization of small GTPase Rab1a as a target of an <i>N</i>-arylbenzimidazole ligand's rescue of Parkinson's-associated cell toxicity

Hatstat, A. Katherine; Quan, Baiyi; Bailey, Morgan A; Fitzgerald, Michael C.; Reinhart, Michaela C.; McCafferty, Dewey G.

doi:10.1039/d1cb00103e

Cited by 8 publications

(5 citation statements)

References 81 publications

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“…Because activation of both Rab1A and Rab1B were implicated in multiple cancer types ( Yang et al, 2016 ), both should be considered when studying cancer. In contrast, only Rab1A, and not Rab1B, has been associated with neurological disorders, for example, Parkinson’s disease ( Winslow et al, 2010 ; Coune et al, 2011 ; Ejlerskov et al, 2013 ; Mazzulli et al, 2016 ; Hatstat et al, 2022 ), Alzheimer’s disease ( Mohamed et al, 2017 ), amyotrophic lateral sclerosis ( Webster et al, 2018 ), and intellectual disability ( Tabata et al, 2022 ). In light of the results presented here, we propose that it is the function of Rab1A in autophagy that is important for its role in neurological disorders.…”

Section: Discussionmentioning

confidence: 99%

Dual function of Rab1A in secretion and autophagy: hypervariable domain dependence

et al. 2023

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We currently understand how the different intracellular pathways, secretion, endocytosis, and autophagy are regulated by small GTPases. In contrast, it is unclear how these pathways are coordinated to ensure efficient cellular response to stress. Rab GTPases localize to specific organelles through their hypervariable domain (HVD) to regulate discrete steps of individual pathways. Here, we explored the dual role of Rab1A/B (92% identity) in secretion and autophagy. We show that although either Rab1A or Rab1B is required for secretion, Rab1A, but not Rab1B, localizes to autophagosomes and is required early in stress-induced autophagy. Moreover, replacing the HVD of Rab1B with that of Rab1A enables Rab1B to localize to autophagosomes and regulate autophagy. Therefore, Rab1A-HVD is required for the dual functionality of a single Rab in two different pathways: secretion and autophagy. In addition to this mechanistic insight, these findings are relevant to human health because both the pathways and Rab1A/B were implicated in diseases ranging from cancer to neurodegeneration.

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Section: Discussionmentioning

confidence: 99%

Dual function of Rab1A in secretion and autophagy: hypervariable domain dependence

et al. 2023

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“… 356 NAB2 (N-arylbenzimidazole) was identified to alleviate α-Synuclein induced toxicity in PD cell models, and a following chemoproteomic study reveals that RAB1 is a target of NAB2 for its rescuing effects. 357 Exploring the allosteric pockets on the Rab protein surfaces has also been pursued based on structure-based computational analysis. 358 , 359 Relative to other families, drug discovery efforts toward Ran are limited, though there have been some early attempts to interfere with RAN-GEF interactions.…”

Section: Targeting Small Gtpases In Human Diseasesmentioning

confidence: 99%

Targeting small GTPases: emerging grasps on previously untamable targets, pioneered by KRAS

Yin

Huang

Petela

et al. 2023

Sig Transduct Target Ther

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Small GTPases including Ras, Rho, Rab, Arf, and Ran are omnipresent molecular switches in regulating key cellular functions. Their dysregulation is a therapeutic target for tumors, neurodegeneration, cardiomyopathies, and infection. However, small GTPases have been historically recognized as “undruggable”. Targeting KRAS, one of the most frequently mutated oncogenes, has only come into reality in the last decade due to the development of breakthrough strategies such as fragment-based screening, covalent ligands, macromolecule inhibitors, and PROTACs. Two KRASG12C covalent inhibitors have obtained accelerated approval for treating KRASG12C mutant lung cancer, and allele-specific hotspot mutations on G12D/S/R have been demonstrated as viable targets. New methods of targeting KRAS are quickly evolving, including transcription, immunogenic neoepitopes, and combinatory targeting with immunotherapy. Nevertheless, the vast majority of small GTPases and hotspot mutations remain elusive, and clinical resistance to G12C inhibitors poses new challenges. In this article, we summarize diversified biological functions, shared structural properties, and complex regulatory mechanisms of small GTPases and their relationships with human diseases. Furthermore, we review the status of drug discovery for targeting small GTPases and the most recent strategic progress focused on targeting KRAS. The discovery of new regulatory mechanisms and development of targeting approaches will together promote drug discovery for small GTPases.

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“…The TPP, CPP, SPROX, PP, LiP, and DART techniques have been most widely used in protein target discovery efforts where the goal is to identify the protein targets of a test ligand. An increasing number of successes using these methods to identify protein targets of drugs ,− and other biologically relevant molecules ,,− have been reported by us and others. In these protein target discovery studies, there is typically much overlap in the protein targets identified using each technique, and it has been noted in some reports that using a combination of these strategies can help differentiate true positives from false positives. ,,, A recent comparative analysis of the analytical capabilities of SPROX, TPP, CPP, and PP using the well-studied ligand, cyclosporine A (CSA), and yeast cell lysate showed the benefits of using all four strategies, the biggest one of which is that the false positive rate can be reduced from 30 to 70% to near 0% if selected protein hits are required to appear in at least two different techniques.…”

Section: Introductionmentioning

confidence: 99%

“…In these protein target discovery studies, there is typically much overlap in the protein targets identified using each technique, and it has been noted in some reports that using a combination of these strategies can help differentiate true positives from false positives. 14,24,28,29 A recent comparative analysis of the analytical capabilities of SPROX, TPP, CPP, and PP using the well-studied ligand, cyclosporine A (CSA), and yeast cell lysate showed the benefits of using all four strategies, 28 the biggest one of which is that the false positive rate can be reduced from 30 to 70% to near 0% if selected protein hits are required to appear in at least two different techniques.…”

Section: ■ Introductionmentioning

confidence: 99%

Comparative Analysis of Protein Folding Stability-Based Profiling Methods for Characterization of Biological Phenotypes

Bailey

Tang

Park

et al. 2023

J. Am. Soc. Mass Spectrom.

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Recently, a new suite of mass spectrometry-based proteomic methods has been developed that enables evaluation of protein folding stability on the proteomic scale. These methods utilize chemical and thermal denaturation approaches (SPROX and TPP, respectively) as well as proteolysis strategies (DARTS, LiP, and PP) to assess protein folding stability. The analytical capabilities of these technique have been well-established for protein target discovery applications. However, less is known about the relative advantages and disadvantages of using these different strategies to characterize biological phenotypes. Reported here is a comparative study of SPROX, TPP, LiP, and conventional protein expression level measurements using both a mouse model of aging and a mammalian cell culture model of breast cancer. Analyses on proteins in brain tissue cell lysates derived from 1- and 18-month-old mice (n = 4–5 at each time point) and on proteins in cell lysates derived from the MCF-7 and MCF-10A cell lines revealed a majority of the differentially stabilized protein hits in each phenotype analysis had unchanged expression levels. In both phenotype analyses, TPP generated the largest number and fraction of differentially stabilized protein hits. Only a quarter of all the protein hits identified in each phenotype analysis had a differential stability that was detected using multiple techniques. This work also reports the first peptide-level analysis of TPP data, which was required for the correct interpretation of the phenotype analyses performed here. Studies on selected protein stability hits also uncovered phenotype-related functional changes.

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Chemoproteomic-enabled characterization of small GTPase Rab1a as a target of an N-arylbenzimidazole ligand's rescue of Parkinson's-associated cell toxicity

Abstract: The development of phenotypic models of Parkinson's disease (PD) has enabled screening and identification of phenotypically active small molecules that restore complex biological pathways affected by PD toxicity.

Cited by 8 publications

References 81 publications

Dual function of Rab1A in secretion and autophagy: hypervariable domain dependence

Dual function of Rab1A in secretion and autophagy: hypervariable domain dependence

Targeting small GTPases: emerging grasps on previously untamable targets, pioneered by KRAS

Comparative Analysis of Protein Folding Stability-Based Profiling Methods for Characterization of Biological Phenotypes

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