TY - JOUR
T1 - Top-Down, Knowledge-Based Genetic Reduction of Yeast Central Carbon Metabolism
AU - Postma, Eline D.
AU - Couwenberg, Lucas G.F.
AU - van Roosmalen, Roderick N.
AU - Geelhoed, Jordi
AU - de Groot, Philip A.
AU - Daran-Lapujade, Pascale
PY - 2022
Y1 - 2022
N2 - Saccharomyces cerevisiae, whose evolutionary past includes a whole-genome duplication event, is characterized by a mosaic genome configuration with substantial apparent genetic redundancy. This apparent redundancy raises questions about the evolutionary driving force for genomic fixation of “minor” paralogs and complicates modular and combinatorial metabolic engineering strategies. While isoenzymes might be important in specific environments, they could be dispensable in controlled laboratory or industrial contexts. The present study explores the extent to which the genetic complexity of the central carbon metabolism (CCM) in S. cerevisiae, here defined as the combination of glycolysis, the pentose phosphate pathway, the tricarboxylic acid cycle, and a limited number of related pathways and reactions, can be reduced by elimination of (iso)enzymes without major negative impacts on strain physiology. Cas9-mediated, groupwise deletion of 35 of the 111 genes yielded a “minimal CCM” strain which, despite the elimination of 32% of CCM-related proteins, showed only a minimal change in phenotype on glucose-containing synthetic medium in controlled bioreactor cultures relative to a congenic reference strain. Analysis under a wide range of other growth and stress conditions revealed remarkably few phenotypic changes from the reduction of genetic complexity. Still, a well-documented context-dependent role of GPD1 in osmotolerance was confirmed. The minimal CCM strain provides a model system for further research into genetic redundancy of yeast genes and a platform for strategies aimed at large-scale, combinatorial remodeling of yeast CCM.
AB - Saccharomyces cerevisiae, whose evolutionary past includes a whole-genome duplication event, is characterized by a mosaic genome configuration with substantial apparent genetic redundancy. This apparent redundancy raises questions about the evolutionary driving force for genomic fixation of “minor” paralogs and complicates modular and combinatorial metabolic engineering strategies. While isoenzymes might be important in specific environments, they could be dispensable in controlled laboratory or industrial contexts. The present study explores the extent to which the genetic complexity of the central carbon metabolism (CCM) in S. cerevisiae, here defined as the combination of glycolysis, the pentose phosphate pathway, the tricarboxylic acid cycle, and a limited number of related pathways and reactions, can be reduced by elimination of (iso)enzymes without major negative impacts on strain physiology. Cas9-mediated, groupwise deletion of 35 of the 111 genes yielded a “minimal CCM” strain which, despite the elimination of 32% of CCM-related proteins, showed only a minimal change in phenotype on glucose-containing synthetic medium in controlled bioreactor cultures relative to a congenic reference strain. Analysis under a wide range of other growth and stress conditions revealed remarkably few phenotypic changes from the reduction of genetic complexity. Still, a well-documented context-dependent role of GPD1 in osmotolerance was confirmed. The minimal CCM strain provides a model system for further research into genetic redundancy of yeast genes and a platform for strategies aimed at large-scale, combinatorial remodeling of yeast CCM.
KW - central carbon metabolism
KW - genetic redundancy
KW - minimal genome
KW - Saccharomyces cerevisiae
UR - http://www.scopus.com/inward/record.url?scp=85140856828&partnerID=8YFLogxK
U2 - 10.1128/mbio.02970-21
DO - 10.1128/mbio.02970-21
M3 - Article
C2 - 36129294
AN - SCOPUS:85140856828
SN - 2150-7511
VL - 13
SP - e0297021
JO - mBio
JF - mBio
IS - 5
ER -