Dual regulation of the levels and function of Start transcriptional repressors drives G1 arrest in response to cell wall stress

Cell Communication and Signaling, Год журнала: 2025, Номер 23(1)

Опубликована: Янв. 16, 2025

Many different stress signaling pathways converge in a common response: slowdown or arrest cell cycle the G1 phase. The G1/S transition (called Start budding yeast) is key checkpoint controlled by positive and negative regulators. Among them, Whi7 Whi5 are transcriptional repressors of program, yeast functional homologs Retinoblastoma family proteins mammalian cells. Under standard conditions, plays lesser role than inhibition. However, under wall stress, induced more important control. In this work, we investigated hallmarks Whi5, which determine their strength as inhibitors stress. response Saccharomyces cerevisiae to Calcofluor White was characterize regulation function To control protein levels, used dose-dependent β-estradiol-induced expression auxin-induced degron fusions. We also performed Chromatin Immunoprecipitation assays investigate association with promoters scored re-entry using microscopy assays. found that promoted specific upregulation repressor. First, although increases not only determinant behind promoting arrest. Indeed, artificial induction at same level resulted lower block. Second, specifically recruited SBF-target promoters, independent increase its levels stage. Finally, instability further increased during degradation triggered advanced re-entry. Here, show enhances Importantly, identified new Whi7-specific regulatory mechanisms do operate Our results indicate cells may benefit from stress-specific ensure stress-induced rapid be particularly resume upon adaptation.

Язык: Английский

Deep learning-driven imaging of cell division and cell growth across an entire eukaryotic life cycle

bioRxiv (Cold Spring Harbor Laboratory), Год журнала: 2024, Номер unknown

Опубликована: Апрель 27, 2024

Abstract The life cycle of biomedical and agriculturally relevant eukaryotic microorganisms involves complex transitions between proliferative non-proliferative states such as dormancy, mating, meiosis, cell division. New drugs, pesticides, vaccines can be created by targeting specific stages parasites pathogens. However, defining the structure a microbial often relies on partial observations that are theoretically assembled in an ideal path. To create more quantitative approach to studying complete cycles, we generated deep learning-driven imaging framework track across sexually reproducing generations. Our combines microfluidic culturing, stage-specific segmentation microscopy images using convolutional neural networks, novel tracking algorithm, FIEST, based enhancing overlap single masks consecutive through learning video frame interpolation. As proof principle, used this quantitatively image compare growth regulation sexual Saccharomyces cerevisiae . We developed fluorescent reporter system fluorescently labeled Whi5 protein, yeast analog mammalian Rb, new High-Cdk1 activity sensor, LiCHI, designed report during DNA replication, mitosis, meiotic homologous recombination, meiosis I, II. found preceded exit from mitotic G1, pre-meiotic G0 spore state germination. A decrease total concentration characterized states, which is consistent with dilution model. nuclear accumulation was developmentally regulated, being at its highest formation. temporal coordination division not significantly different three could characterize other single-cell cycles remain incompletely described. An off-the-shelf user interface Yeastvision provides free access our processing algorithms.

Язык: Английский

A non-canonical CDK, Pho85 regulates the restart of the cell-cycle following stress

bioRxiv (Cold Spring Harbor Laboratory), Год журнала: 2024, Номер unknown

Опубликована: Авг. 27, 2024

Abstract Environmental stress induces an arrest of the cell cycle. Thus, release from this is essential for survival. The cell-cycle-arrest occurs via down regulation cyclins that drive main cyclin dependent kinase, CDK1/Cdc28. However, it was not clear how cells escape potentially fatal arrest. Here we show prior to restoration CDK1/Cdc28 cyclins, a non-canonical CDK, Pho85, initiates cascade restart We demonstrate following stress, Pho85 phosphorylates Sch9 which in turn directly transcriptional inhibitor Whi5, yeast analog RB1/retinoblastoma, and CDK1 target. This promotes Whi5 translocation nucleus, stress-induced at G 1 phase. In addition, find parallel with also plays role control Whi5. Together, these findings provide insights into re-enter cycle during recovery reveal CDK takes on roles acts pathway functions

Язык: Английский