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Gene Expression Regulates Metabolite Homeostasis During the Crabtree Effect: Implications for the Adaptation and Evolution of Metabolism

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Gene Expression Regulates Metabolite Homeostasis During the Crabtree Effect: Implications for the Adaptation and Evolution of Metabolism Gene expression regulates metabolite homeostasis during the Crabtree effect: Implications for the adaptation and evolution of Metabolism Douglas L. Rothmana,b,1, Stephen C. Stearnsc, and Robert G. Shulmanb,1 aDepartment of Radiology & Biomedical Engineering, Yale University, New Haven, CT 06520; bMagnetic Resonance Research Center, Yale University, New Haven, CT 06520; and cDepartment of Ecology and Evolutionary Biology, Yale University, New Haven, CT 06520 Contributed by Robert G. Shulman, November 24, 2020 (sent for review July 6, 2020; reviewed by Barbara M. Bakker and Jan-Hendrik S. Hofmeyr) A key issue in both molecular and evolutionary biology has been to hours; in evolutionary biology, the genetic changes involve mu- to define the roles of genes and phenotypes in the adaptation of tations, changes in gene frequency, gene duplications, and the organisms to environmental changes. The dominant view has been rewiring, via those mechanisms, of genetic control networks, all that an organism’s metabolic adaptations are driven by gene expres- processes that take many generations. In both cases, plasticity pre- sion and that gene mutations, independent of the starting phenotype, cedes genetic change. These nuanced interactions are the subject of are responsible for the evolution of new metabolic phenotypes. We this paper. propose an alternate hypothesis, in which the phenotype and geno- The genome centric view of metabolic adaptation originated type together determine metabolic adaptation both in the lifetime of with Beadle (7), who introduced the concept, often summarized the organism and in the evolutionary selection of adaptive metabolic as “one gene, one enzyme,” and used it to explain in born errors traits. We tested this hypothesis by flux-balance and metabolic-control of metabolism. This concept was extended to coordinated ex- analysis of the relative roles of the starting phenotype and gene ex- pression of multiple genes in the pioneering work of Jacob and pression in regulating the metabolic adaptations during the Crabtree Monod (8), who showed in bacteria that the genes coding the effect in yeast, when they are switched from a low- to high-glucose enzymes in the lactose synthesis pathway are transcribed at an environment. Critical for successful short-term adaptation was the equal rate due to being on the same stretch of DNA that is under ability of the glycogen/trehalose shunt to balance the glycolytic path- control of a common repressor element that binds RNA poly- BIOCHEMISTRY way. The role of later gene expression of new isoforms of glycolytic merase. In the absence of lactose, the polymerase is blocked enzymes, rather than flux control, was to provide additional homeo- by the repressor protein. When lactose in the environment static mechanisms allowing an increase in the amount and efficiency increases, the repressor protein dissociates, allowing the poly- of adenosine triphosphate and product formation while maintaining merase to coordinately express the pathway enzymes, which re- glycolytic balance. We further showed that homeostatic mechanisms, sults in a proportional increase in the lactose-consumption rate. by allowing increased phenotypic plasticity, could have played an im- They referred to this mechanism as an operon and described it as portant role in guiding the evolution of the Crabtree effect. Although a “genetic regulatory mechanism” (8). These and subsequent our findings are specific to Crabtree yeast, they are likely to be broadly findings led to the view that metabolic regulation and the found because of the well-recognized similarities in glucose metabo- lism across kingdoms and phyla from yeast to humans. Significance glucose metabolism | glycogen shunt | adaptation | homeostasis | Crabtree effect The dominant view in both molecular and evolutionary biology is that genotype controls the adaptation to new environments. or more than a 100 y, a central goal of biology has been to We propose an alternate hypothesis, in which the phenotype Fdetermine the roles that genes and phenotypes play in the and genotype both play important roles in metabolic adapta- adaptation of an organism to a change in the environment. In tion in the lifetime of the organism and in the evolutionary molecular biology, the dominant paradigm has been top-down selection of adaptive metabolic traits. When studying the genomic determination of phenotypes, including the pathogen- Crabtree effect in yeast, we found that homeostatic mecha- esis of disease, a major motivation for the human genome project nisms in the starting phenotype, primarily the glycogen/tre- (1). However, this viewpoint has been challenged, both due to halose shunt, allow adaptation to high glucose. Subsequent the increasing difficulty of mapping DNA elements to genes and gene expression, rather than creating a new phenotype, opti- functions (2) and failures to predict phenotypes from genotypes. mizes the initial adaptation by expression of new homeostatic McKnight (3), reviewing studies of cancer metabolism, proposed mechanisms. Metabolic homeostatic mechanisms, by allowing that, due to feedback, a strict top-down genomic control model increased phenotypic plasticity, could also have played an was not sufficient and that direct studies of the metabolic phe- important role in guiding the evolution of the Crabtree notype were necessary to understand metabolic adaptation. In phenotype. evolutionary biology, the Modern Synthesis of the mid-20th Author contributions: D.L.R., S.C.S., and R.G.S. designed research; D.L.R., S.C.S., and R.G.S. century was also a paradigm that gave genes a dominant role; performed research; D.L.R., S.C.S., and R.G.S. analyzed data; and D.L.R., S.C.S., and R.G.S. it used changes in gene frequencies over many generations to wrote the paper. explain evolutionary adaptations (4, 5). That view of allelic Reviewers: B.M.B., University Medical Center Groningen; and J.-H.S.H., Stellenbosch substitutions leading directly to the development of new phe- University. notypes has been modified by the proposal that phenotypic The authors declare no competing interest. plasticity, defined as the range of phenotypes that can be sup- Published under the PNAS license. ported by a genotype (and its population variations), can extend 1To whom correspondence may be addressed. Email: [email protected] or the range of novel environments in which organisms can survive, [email protected]. allowing time for selection to supplement more rapid plastic re- This article contains supporting information online at https://www.pnas.org/lookup/suppl/ sponses with slower genetic changes (5, 6). In metabolism, the doi:10.1073/pnas.2014013118/-/DCSupplemental. genetic changes involve gene expression, which takes minutes Published December 28, 2020. PNAS 2021 Vol. 118 No. 2 e2014013118 https://doi.org/10.1073/pnas.2014013118 | 1of11 Downloaded by guest on September 27, 2021 evolution of new metabolic phenotypes can be understood pri- metabolism soon after glucose exposure. It is one of many ex- marily from genomic analysis (1). amples in unicellular organisms (22) and multicellular organisms Though conceptually powerful, relatively simple operon mech- (16–19) in which metabolic pathways can respond rapidly to a anisms have not been found in eukaryotes (2). However, findings change in environment prior to a change in gene expression. Al- of coordinated gene expression, initially in yeast exposed to high though specific to yeast, it is relevant to a wide range of organisms, glucose (9), have provided continuing—albeit indirect—support including mammals, for it involves the coregulation of the ubiq- for top-down genomic regulation of metabolic adaptation. How- uitous glycolytic, gluconeogenic, and glycogen-synthesis pathways ever, this conclusion has been challenged by studies using quan- that play a central role in glucose metabolism. titative methods such as metabolic control analysis (10), which When Crabtree yeast are first exposed to glucose, their glycolytic found that gene expression often does not have a major role in the pathwayisinanimbalancedstateinwhichtherearehighactivities adaptation of metabolic flux to environmental changes. Rather, of the enzymes that supply glucose to the glycolytic pathway, such as the flux values after adaptation are often primarily determined by glucose transport (GT), hexokinase (HK), and phosphofructokinase nongenomic mechanisms, such as allosteric activation and inhibi- (PFK), but reduced enzyme activity for the lower demand portion tion of enzymes (11, 12), and posttranslational enzyme modifica- of glycolysis that couples the flux to cellular ATP needs, in partic- tions (13), such as phosphorylation (14, 15). Similar conclusions ular, pyruvate kinase (PK) (Fig. 1B, “Early Fermentation State”). have been reached regarding mammalian glucose and mitochon- The initial relatively low enzyme activity of the demand portion of drial metabolism (16–19). However, while in these systems, the the pathway is important during the respiratory state, because it primary adaptations in flux are driven by the initial metabolic allows glucose 6-phosphate (G6P) production by gluconeogenesis phenotype, the accompanying widespread changes in gene ex- for the pentose phosphate pathway (23). However, when there is a pression remain largely unexplained. major influx of glucose into the cell, the resultant imbalance, if not To address the interplay of genomic and nongenomic mech- compensated, leads to large increases of fructose bisphosphate
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