Muralidhara, P. | bioRxiv Science

bioRxiv2021

When Yeast Cells Change their Mind: Cell Cycle 'Start' is Reversible under Starvation

Eukaryotic cells decide in late G1 whether to commit to another round of genome duplication and division. This point of irreversible cell cycle commitment is a molecular switch termed "Restriction Point" in mammals and "Start" in budding yeast. At Start, yeast cells integrate multiple signals such as pheromones, osmolarity, and nutrients. If sufficient nutrients are lacking, cells will not pass Start. However, how the cells respond to nutrient depletion after they have made the Start decision, remains poorly understood. Here, we analyze by live cell imaging how post-Start yeast cells respond to nutrient depletion. We monitor fluorescently labelled Whi5, the cell cycle inhibitor whose export from the nucleus determines Start. Surprisingly, we find that cells that have passed Start can re-import Whi5 back into the nucleus. This occurs when cells are faced with starvation up to 20 minutes after Start. In these cells, the positive feedback loop is interrupted, Whi5 re-binds DNA, and CDK activation occurs a second time once nutrients are replenished. Cells which re-import Whi5 also become sensitive to mating pheromone again, and thus behave like pre-Start cells. In summary, we show that upon starvation the commitment decision at Start can be reversed. We therefore propose that in yeast, as has been suggested for mammalian cells, cell cycle commitment is a multi-step process, where irreversibility in face of nutrient signaling is only reached approximately 20 minutes after CDK activation at Start.

cell biology↗

bioRxiv2020

Global transcriptional regulation by cell-free supernatant of Salmonella Typhimurium peptide transporter mutant leads to inhibition of intra-species biofilm initiation

Salmonella is a genus of widely spread Gram negative, facultative anaerobic bacteria, which is known to cause [1/4]th of diarrheal morbidity and mortality globally. It causes typhoid fever and gastroenteritis by gaining access to the host gut through contaminated food and water. Salmonella utilizes its biofilm lifestyle to strongly resist antibiotics and persist in the host. Although biofilm removal or dispersal has been studied widely, the inhibition of the initiation of Salmonella biofilm remains elusive. This study was conducted to determine the anti-biofilm property of the cell-free supernatant obtained from a carbon-starvation inducible proline peptide transporter mutant ({Delta}yjiY) strain. Our study shows that Salmonella {Delta}yjiY culture supernatant primarily inhibits biofilm initiation by regulating biofilm-associated transcriptional network. This work demonstrates that highly abundant proteases such as HslV and GrpE cleave the protein aggregates, whereas global transcription regulators H-NS, FlgM regulate expression of SPIs and flagellar genes. Relatively low abundances of flavoredoxin, glutaredoxin, thiol peroxidase etc. leads to accumulation of ROS within the biofilm, and subsequent toxicity. This work further suggests that targeting these oxidative stress relieving proteins might be a good druggable choice to reduce Salmonella biofilm. ImportanceThe enteric pathogen Salmonella forms biofilm in the internal organs of asymptomatic carriers, and on abiotic surfaces that leads to contamination of food and water. Biofilms are highly drug-resistant life forms that also helps in host immune evasion. Therefore, recent insurgence of drug tolerant strains necessitates development of biofilm inhibitory strategies, and finding novel druggable targets. In this study we investigated the bioactive molecules present in the cell-free supernatant of a biofilm deficient strain of Salmonella Typhimurium that inhibit biofilm initiation by the wildtype strain. Further we showed that the supernatant treatment leads to virulence defect in vivo. Collectively, our results suggest a comprehensive view of virulence regulation in Salmonella by the cell-free supernatant of the biofilm deficient strain.

microbiology↗