High-resolution yeast quiescence profiling in human-like media reveals complex influences of auxotrophy and nutrient ava
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ORIGINAL ARTICLE
High-resolution yeast quiescence profiling in human-like media reveals complex influences of auxotrophy and nutrient availability Sean M. Santos & Samantha Laflin & Audrie Broadway & Cosby Burnet & Joline Hartheimer & John Rodgers & Daniel L. Smith Jr & John L. Hartman IV Received: 20 May 2020 / Accepted: 3 September 2020 # American Aging Association 2020
Abstract Yeast cells survive in stationary phase culture by entering quiescence, which is measured by colonyforming capacity upon nutrient re-exposure. Yeast chronological lifespan (CLS) studies, employing the comprehensive collection of gene knockout strains, have correlated weakly between independent laboratories, which is hypothesized to reflect differential interaction between the deleted genes, auxotrophy, media composition, and other assay conditions influencing quiescence. This hypothesis was investigated by highthroughput quiescence profiling of the parental prototrophic strain, from which the gene deletion strain libraries were constructed, and all possible auxotrophic allele combinations in that background. Defined media resembling human cell culture media promoted longterm quiescence and was used to assess effects of glucose, ammonium sulfate, auxotrophic nutrient availability, target of rapamycin signaling, and replication stress. Frequent, high-replicate measurements of colonyforming capacity from cultures aged past 60 days provided profiles of quiescence phenomena such as gasping and hormesis. Media acidification was assayed in parallel to assess correlation. Influences of leucine, Electronic supplementary material The online version of this article (https://doi.org/10.1007/s11357-020-00265-2) contains supplementary material, which is available to authorized users. S. M. Santos : S. Laflin : A. Broadway : C. Burnet : J. Hartheimer : J. Rodgers : D. L. Smith Jr : J. L. Hartman IV (*) Department of Genetics, University of Alabama at Birmingham, Birmingham, AL, USA e-mail: [email protected]
methionine, glucose, and ammonium sulfate metabolism were clarified, and a role for lysine metabolism newly characterized, while histidine and uracil perturbations had less impact. Interactions occurred between glucose, ammonium sulfate, auxotrophy, auxotrophic nutrient limitation, aeration, TOR signaling, and/or replication stress. Weak correlation existed between media acidification and maintenance of quiescence. In summary, experimental factors, uncontrolled across previous genome-wide yeast CLS studies, influence quiescence and interact extensively, revealing quiescence as a complex metabolic and developmental process that should be studied in a prototrophic context, omitting ammonium sulfate from defined media, and employing highly replicable protocols. Keywords Quantitative high-throughput cell array phenotyping (Q-HTCP) . Quiescence profiling . Yeast chronological lifespan(CLS) . Gene-nutrient interaction . Human-like (HL) yeast media
Introduction Cell quiescence is the process of adaptive cell cycle exit (G0), prior to the G1-S transition,
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