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Fast growth increases the selective advantage of a mutation arising recurrently during evolution under metal limitation.

ABSTRACT: Understanding the evolution of biological systems requires untangling the molecular mechanisms that connect genetic and environmental variations to their physiological consequences. Metal limitation across many environments, ranging from pathogens in the human body to phytoplankton in the oceans, imposes strong selection for improved metal acquisition systems. In this study, we uncovered the genetic and physiological basis of adaptation to metal limitation using experimental populations of Methylobacterium extorquens AM1 evolved in metal-deficient growth media. We identified a transposition mutation arising recurrently in 30 of 32 independent populations that utilized methanol as a carbon source, but not in any of the 8 that utilized only succinate. These parallel insertion events increased expression of a novel transporter system that enhanced cobalt uptake. Such ability ensured the production of vitamin B(12), a cobalt-containing cofactor, to sustain two vitamin B(12)-dependent enzymatic reactions essential to methanol, but not succinate, metabolism. Interestingly, this mutation provided higher selective advantages under genetic backgrounds or incubation temperatures that permit faster growth, indicating growth-rate-dependent epistatic and genotype-by-environment interactions. Our results link beneficial mutations emerging in a metal-limiting environment to their physiological basis in carbon metabolism, suggest that certain molecular features may promote the emergence of parallel mutations, and indicate that the selective advantages of some mutations depend generically upon changes in growth rate that can stem from either genetic or environmental influences.

SUBMITTER: Chou HH

PROVIDER: S-EPMC2732905 | biostudies-literature | 2009 Sep

REPOSITORIES: biostudies-literature

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Fast growth increases the selective advantage of a mutation arising recurrently during evolution under metal limitation.

Chou Hsin-Hung HH Berthet Julia J Marx Christopher J CJ

PLoS genetics 20090918 9

Understanding the evolution of biological systems requires untangling the molecular mechanisms that connect genetic and environmental variations to their physiological consequences. Metal limitation across many environments, ranging from pathogens in the human body to phytoplankton in the oceans, imposes strong selection for improved metal acquisition systems. In this study, we uncovered the genetic and physiological basis of adaptation to metal limitation using experimental populations of Methy ...[more]

PMID: 19763169

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Project description:BACKGROUND:Adaptive responses to nutrient limitation involve mutations that increase the efficiency of usage or uptake of the limiting nutrient. However, starvation of different nutrients has contrasting effects on physiology, resulting in different evolutionary responses. Most studies performed to understand these evolutionary responses have focused only on macronutrient limitation. Hence our understanding of adaptation under limitation of other forms of nutrients is limited. In this study, we compared the evolutionary response in populations evolving under growth-limiting conditions for a macronutrient and a major cation. RESULTS:We evolved eight populations of E. coli in nutrient-limited chemostats for 400 generations to identify the genetic basis of the mechanisms involved in efficient usage of two nutrients: nitrogen and magnesium. Our population genomic sequencing work, based on this study and previous work, allowed us to identify targets of selection under these nutrient limiting conditions. Global transcriptional regulators glnGL were targets of selection under nitrogen starvation, while proteins involved in outer-membrane biogenesis (genes from the lpt operon) were targets of selection under magnesium starvation. The protein involved in cell-cycle arrest (yhaV) was a target of selection in both environments. We re-constructed specific mutants to analyze the effect of individual mutations on fitness in nutrient limiting conditions in chemostats and in batch cultures. We further demonstrated that adaptation to nitrogen starvation proceeds via a nutrient specific mechanism, while that to magnesium starvation involves a more general mechanism. CONCLUSIONS:Our results show two different forms of adaptive strategies under limitation of nutrients that effect cellular physiology in different ways. Adaptation to nitrogen starvation proceeds by upregulation of transcriptional regulator glnG and subsequently of transporter protein amtB, both of which results in increased nitrogen scavenging ability of the cell. On the other hand, adaptation to magnesium starvation proceeds via the restructuring of the cell outer-membrane, allowing magnesium to be redistributed to other biological processes. Also, adaptation to the chemostat environment involves selection for loss of function mutations in genes that under nutrient-limiting conditions interfere with continuous growth.

Project description:BACKGROUND:Glucoamylase is one of the most industrially applied enzymes, produced by Aspergillus species, like Aspergillus niger. Compared to the traditional ways of process optimization, the metabolic engineering strategies to improve glucoamylase production are relatively scarce. RESULTS:In the previous study combined multi-omics integrative analysis and amino acid supplementation experiment, we predicted four amino acids (alanine, glutamate, glycine and aspartate) as the limited precursors for glucoamylase production in A. niger. To further verify this, five mutants namely OE-ala, OE-glu, OE-gly, OE-asp1 and OE-asp2, derived from the parental strain A. niger CBS 513.88, were constructed respectively for the overexpression of five genes responsible for the biosynthesis of the four kinds of amino acids (An11g02620, An04g00990, An05g00410, An04g06380 and An16g05570). Real-time quantitative PCR revealed that all these genes were successfully overexpressed at the mRNA level while the five mutants exhibited different performance in glucoamylase production in shake flask cultivation. Notably, the results demonstrated that mutant OE-asp2 which was constructed for reinforcing cytosolic aspartate synthetic pathway, exhibited significantly increased glucoamylase activity by 23.5% and 60.3% compared to CBS 513.88 in the cultivation of shake flask and the 5 L fermentor, respectively. Compared to A. niger CBS 513.88, mutant OE-asp2 has a higher intracellular amino acid pool, in particular, alanine, leucine, glycine and glutamine, while the pool of glutamate was decreased. CONCLUSION:Our study combines the target prediction from multi-omics analysis with the experimental validation and proves the possibility of increasing glucoamylase production by enhancing limited amino acid biosynthesis. In short, this systematically conducted study will surely deepen the understanding of resources allocation in cell factory and provide new strategies for the rational design of enzyme production strains.

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Fast growth increases the selective advantage of a mutation arising recurrently during evolution under metal limitation.

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Fast growth increases the selective advantage of a mutation arising recurrently during evolution under metal limitation.

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