The Role of mRNA Degradation in Dynamic Nitrogen Environments in Saccharomyces cerevisiae

Abstract

Studying the reprogramming of gene expression is fundamental to understanding cellular growth, development and stem cell differentiation. In Saccharomyces cerevisiae cells undergo reprogramming of gene expression when they are grown in nitrogen‐poor conditions and then switched to nitrogen‐rich conditions. This is controlled by the nitrogen catabolite repression (NCR) response. In a previous study, NCR transcripts were shown to undergo accelerated degradation when the cells where switched from growth on proline in a nitrogen‐poor environment to growth in a nitrogen‐rich environment upon the addition of glutamine. This study aims to determine the benefit of accelerated mRNA degradation in dynamic nitrogen environments by studying the effects of mutants of NCR repressors during this transition. We suspect that knocking out the repressors will stop the accelerated degradation once glutamine is added. This will in turn affect the growth rates and cause the mutants to have longer lag phase, a period where there is no detectable change in the growth. The general amino acid permease (GAP1) was used in representation of NCR transcripts to study the effects. This gene encodes a permease that senses the presence of amino acids substrates to regulate localization to the plasma membrane when needed. We identified mutants where GAP1 is stabilized in a nitrogen‐rich environment by using knock‐outs (KO) of the NCR repressor genes Ure2, Dal80 and Gzf3. The Ure2KO mutant showed depression of GAP1 in nitrogen‐rich conditions, but it caused a growth defect when the cells were grown in nitrogen‐poor conditions. In addition, we tested the effect of the GAP1 5′ untranslated region (UTR) mutant on growth dynamics because it has been shown to stabilize the GAP1 transcript during the transition from nitrogen‐poor environment to a nitrogen‐rich environment. A T‐test was performed to see if there was any difference between the growth rates of the wildtype and the 5′UTR deletion. Turns out that there was no significant difference between them before (p value=0.13) and after the glutamine addition (p value= 0.15), despite there being a notable distinction between the slopes. When the graphs for the nitrogen up shift were generated a lag phase was detected and it appeared to be an hour longer in the mutant. Which indicates that accelerated degradation is an important process in dynamic nitrogen environments.Support or Funding InformationNIH‐MARC Program undergraduate fellowship (5T34GM00781‐37)