A natural variant of the sole pyruvate kinase of fission yeast lowers glycolytic flux triggering increased respiration and oxidative-stress resistance but decreased growth

Abstract

Cells balance glycolysis with respiration to support their energetic and biosynthetic needs in different environmental or physiological contexts. With abundant glucose, many cells prefer to grow by aerobic glycolysis, or fermentation in yeast. Using 161 natural isolates of fission yeast, we investigated the genetic basis and phenotypic effects of the fermentation-respiration balance. The laboratory and a few other strains were more dependent on respiration. This trait was associated with a missense variant in a highly conserved region of Pyk1. Pyk1 is the single pyruvate kinase in fission yeast, while most organisms possess isoforms with different activity. This variant reduced Pyk1 activity and glycolytic flux. Replacing the 'low-activity' pyk1 allele in the laboratory strain with the common 'high-activity' allele was sufficient to increase fermentation and decrease respiration. This metabolic reprogramming triggered systems-level adaptations in the transcriptome and proteome, and in cellular phenotypes, including increased growth and chronological lifespan, but decreased resistance to oxidative stress. Thus, low Pyk1 activity provided no growth advantage but stress tolerance, despite increased respiration. The genetic tuning of glycolytic flux by a single-nucleotide change might reflect an adaptive trade-off in a species lacking pyruvate-kinase isoforms.

Mitochondrial respiration is required to provide amino acids during fermentative proliferation of fission yeast

“…Some fission yeast isolates, including the standard laboratory strain, possess a mutation in pyruvate kinase, a key glycolytic enzyme [34]. This feature limits glycolytic flux and causes increased respiration during fermentative growth [34]. It is possible that the shortage of amino acids is exacerbated by this mutation.…”

“…However, such adaptations are the exception: most yeast species, including fission yeast (Schizosaccharomyces pombe), are petite-negative and do not cope well under anaerobic conditions. The S. pombe laboratory strain is part of a group of natural isolates that show higher respiration during fermentative growth, which is caused by a naturally occurring mutation in pyruvate kinase that limits glycolytic flux [34]. We have also reported that respiration is required for rapid cell proliferation during fermentation [35].…”

Measuring glycolytic flux in single yeast cells with an orthogonal synthetic biosensor

“…Here, drawing on the glycolytic flux‐signaling metabolite fructose‐1,6‐bisphosphate (FBP) in yeast (Huberts et al , ; Hackett et al , ; preprint: Kamrad et al , ) and using the B. subtilis FBP‐binding transcription factor CggR (Doan & Aymerich, ), we developed a biosensor that allows for sensing FBP levels, and thus glycolytic flux, in single yeast cells. To this end, we used computational protein design, biochemical, proteome, and metabolome analyses (i) to develop a synthetic yeast promoter regulated by the bacterial transcription factor CggR, (ii) to engineer the transcription factor's FBP‐binding site toward increasing the sensor's dynamic range, and (iii) to establish growth‐independent CggR expression levels.…”