Quantitative determinants of aerobic glycolysis identify flux through the enzyme GAPDH as a limiting step

Shestov, Alexander A.; Liu, Xiaojing; Ser, Zheng; Cluntun, Ahmad A.; Hung, Yin P.; Huang, Lei; Kim, Dongsung; Le, Anne; Yellen, Gary; Albeck, John G.; Locasale, Jason W.

doi:10.7554/elife.03342

Cited by 230 publications

(231 citation statements)

References 55 publications

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“…Another quantified cysteine outside of the GAPDH active site – C247 (Figure 1F) – was unaltered in reactivity by NRF2 knockdown (Figure 1F), and we confirmed that GAPDH protein expression was unaffected in shNRF2 cells by immunoblotting (Figure 1H). C152 in GAPDH is a redox-sensitive residue that is subject to S-sulphenylation (Yang et al, 2014) and S-sulfhydration (Shestov et al, 2014) and can be affected by pharmacologically induced forms of oxidative stress (Yun et al, 2015). To our knowledge, however, the regulation of C152 by NRF2 has not been previously reported.…”

Section: Resultsmentioning

confidence: 99%

Chemical Proteomics Identifies Druggable Vulnerabilities in a Genetically Defined Cancer

Bar-Peled

Kemper

Suciu

et al. 2017

Cell

219

223

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The transcription factor NRF2 is a master regulator of the cellular antioxidant response and is often genetically activated in Non-Small Cell Lung Cancers (NSCLCs) by, for instance, mutations in the interacting protein KEAP1. While direct pharmacological inhibition of NRF2 has proven challenging, its aberrant activation rewires biochemical networks in cancer cells that may create special vulnerabilities. Here, we use chemical proteomics to map druggable proteins that are selectively expressed in KEAP1-mutant NSCLC cells. Principal among these was NR0B1, an atypical orphan nuclear receptor that we show engages in a multimeric protein complex to regulate the transcriptional output of KEAP1-mutant NSCLC cells. We further identify small molecules that covalently target a conserved cysteine within the NR0B1 protein interaction domain and demonstrate that these compounds disrupt NR0B1 complexes and impair the anchorage-independent growth of KEAP1-mutant cancer cells. Our findings designate NR0B1 as a druggable, transcriptional regulator that supports NRF2-dependent lung cancers.

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Section: Resultsmentioning

confidence: 99%

Chemical Proteomics Identifies Druggable Vulnerabilities in a Genetically Defined Cancer

Bar-Peled

Kemper

Suciu

et al. 2017

Cell

219

223

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“…In fact, DENV infection upregulates glucose transporter 1 and HK2 (44). Although GAPDH is not classically defined as a rate-limiting enzyme, recent reports have identified it to be a bottleneck in the aerobic glycolysis process because it can be regulated by important glycolytic intermediates, such as ATP and NAD ϩ (45). Together, our finding that DENV infection increases GAPDH activity shows that increased GAPDH activity represents an additional mechanism used by this virus to enhance glycolysis and thereby establish a successful infection.…”

Section: Discussionmentioning

confidence: 99%

Dengue Virus NS1 Protein Modulates Cellular Energy Metabolism by Increasing Glyceraldehyde-3-Phosphate Dehydrogenase Activity

Allonso

Andrade

Conde

et al. 2015

J Virol

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Dengue is one of the main public health concerns worldwide. Recent estimates indicate that over 390 million people are infected annually with the dengue virus (DENV), resulting in thousands of deaths. Among the DENV nonstructural proteins, the NS1 protein is the only one whose function during replication is still unknown. NS1 is a 46-to 55-kDa glycoprotein commonly found as both a membrane-associated homodimer and a soluble hexameric barrel-shaped lipoprotein. Despite its role in the pathogenic process, NS1 is essential for proper RNA accumulation and virus production. In the present study, we identified that glyceraldehyde-3-phosphate dehydrogenase (GAPDH) interacts with intracellular NS1. Molecular docking revealed that this interaction occurs through the hydrophobic protrusion of NS1 and the hydrophobic residues located at the opposite side of the catalytic site. Moreover, addition of purified recombinant NS1 enhanced the glycolytic activity of GAPDH in vitro. Interestingly, we observed that DENV infection promoted the relocalization of GAPDH to the perinuclear region, where NS1 is commonly found. Both DENV infection and expression of NS1 itself resulted in increased GAPDH activity. Our findings indicate that the NS1 protein acts to increase glycolytic flux and, consequently, energy production, which is consistent with the recent finding that DENV induces and requires glycolysis for proper replication. This is the first report to propose that NS1 is an important modulator of cellular energy metabolism. The data presented here provide new insights that may be useful for further drug design and the development of alternative antiviral therapies against DENV. IMPORTANCE Dengue represents a serious public health problem worldwide and is caused by infection with dengue virus (DENV).Estimates indicate that half of the global population is at risk of infection, with almost 400 million cases occurring per year. The NS1 glycoprotein is found in both the intracellular and the extracellular milieus. Despite the fact that NS1 has been commonly associated with DENV pathogenesis, it plays a pivotal but unknown role in the replication process. In an effort to understand the role of intracellular NS1, we demonstrate that glyceraldehyde-3-phosphate dehydrogenase (GAPDH) interacts with NS1. Our results indicate that NS1 increases the glycolytic activity of GAPDH in vitro. Interestingly, the GAPDH activity was increased during DENV infection, and NS1 expression alone was sufficient to enhance intracellular GAPDH activity in BHK-21 cells. Overall, our findings suggest that NS1 is an important modulator of cellular energy metabolism by increasing glycolytic flux. Dengue is one of the major health problems in tropical regions. It is estimated that over 390 million people are infected annually with one of the four dengue virus (DENV) serotypes (1). The absence of both an effective tetravalent vaccine and therapeutic agents worsens the impact of the dengue burden. DENV, the most threatening member of the Flaviviridae family,...

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“…Why proliferating cells, including cancer cells, consume large quantities of glucose only to excrete the majority of this carbon as lactate is a subject of debate (Brand, 1985; Brand et al, 1986; DeBerardinis et al, 2008; Gatenby and Gillies, 2004; Hsu and Sabatini, 2008; Hume et al, 1978; Jiang and Deberardinis, 2012; Koppenol et al, 2011; Lunt and Vander Heiden, 2011; Newsholme et al, 1985; Vander Heiden et al, 2009; Vazquez et al, 2010). One widely held hypothesis is that high glycolytic flux allows intermediates to be shunted into anabolic pathways to support biomass accumulation (Brand, 1985; Chaneton et al, 2012; Faubert et al, 2013; Hsu and Sabatini, 2008; Hume et al, 1978; Jiang and Deberardinis, 2012; Jiang et al, 2011; Lunt and Vander Heiden, 2011; Newsholme et al, 1985; Shestov et al, 2014; Vander Heiden et al, 2009; Wang and Green, 2012). Many proliferating mammalian cells also consume substantial quantities of glutamine, and glutamine is also hypothesized to provide material for biosynthesis (Brand, 1985; Brand et al, 1986; Daye and Wellen, 2012; DeBerardinis et al, 2007; Hsu and Sabatini, 2008; Wang and Green, 2012).…”

Section: Introductionmentioning

confidence: 99%

“…This hypothesis has not been rigorously tested, yet it forms the basis for modeling efforts to understand cancer metabolism (Cascante et al, 2002; Shestov et al, 2014; Shlomi et al, 2011). In Escherichia coli , the sources of cell mass and their fates have been carefully quantified (Roberts et al, 1955), and for prototrophic strains grown in defined minimal media, medium composition constrains the nutrients available to produce new biomass.…”

Section: Introductionmentioning

confidence: 99%

Amino Acids Rather than Glucose Account for the Majority of Cell Mass in Proliferating Mammalian Cells

et al. 2016

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Cells must duplicate their mass in order to proliferate. Glucose and glutamine are the major nutrients consumed by proliferating mammalian cells, but the extent to which these and other nutrients contribute to cell mass is unknown. We quantified the fraction of cell mass derived from different nutrients and find that the majority of carbon mass in cells is derived from other amino acids, which are consumed at much lower rates than glucose and glutamine. While glucose carbon has diverse fates, glutamine contributes most to protein, and this suggests that glutamine’s ability to replenish TCA cycle intermediates (anaplerosis) is primarily used for amino acid biosynthesis. These findings demonstrate that rates of nutrient consumption are indirectly associated with mass accumulation and suggest that high rates of glucose and glutamine consumption support rapid cell proliferation beyond providing carbon for biosynthesis.

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Quantitative determinants of aerobic glycolysis identify flux through the enzyme GAPDH as a limiting step

Cited by 230 publications

References 55 publications

Chemical Proteomics Identifies Druggable Vulnerabilities in a Genetically Defined Cancer

Chemical Proteomics Identifies Druggable Vulnerabilities in a Genetically Defined Cancer

Dengue Virus NS1 Protein Modulates Cellular Energy Metabolism by Increasing Glyceraldehyde-3-Phosphate Dehydrogenase Activity

Amino Acids Rather than Glucose Account for the Majority of Cell Mass in Proliferating Mammalian Cells

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