Genetic interactions derived from high-throughput phenotyping of 7,350 yeast cell cycle mutants

AbstractOver the last 30 years, computational biologists have developed increasingly realistic mathematical models of the regulatory networks controlling the division of eukaryotic cells. These models capture data resulting from two complementary experimental approaches: low-throughput experiments aimed at extensively characterizing the functions of small numbers of genes, and large-scale genetic interaction screens that provide a systems-level perspective on the cell division process. The former is insufficient to capture the interconnectivity of the genetic control network, while the latter is fraught with irreproducibility issues. Here, we describe a hybrid approach in which the genetic interactions between 36 cell-cycle genes are quantitatively estimated by high-throughput phenotyping with an unprecedented number of biological replicates. Using this approach, we identify a subset of high-confidence genetic interactions, which we use to refine a previously published mathematical model of the cell cycle. We also present a quantitative dataset of the growth rate of these mutants under six different media conditions in order to inform future cell cycle models.Author SummaryThe process of cell division, also called the cell cycle, is controlled by a highly complex network of interconnected genes. If this process goes awry, diseases such as cancer can result. In order to unravel the complex interactions within the cell cycle control network, computational biologists have developed mathematical models that describe how different cell cycle genes are related. These models are built using large datasets describing the effect of mutating one or more genes within the network. In this manuscript, we present a novel method for producing such datasets. Using our method, we generate 7,350 yeast mutants to explore the interactions between key cell cycle genes. We measure the effect of the mutations by monitoring the growth rate of the yeast mutants under different environmental conditions. We use our mutants to revise an existing model of the yeast cell cycle and present a dataset of ∼44,000 gene by environment combinations as a resource to the yeast genetics and modeling communities.

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Genetic interactions derived from high-throughput phenotyping of 6589 yeast cell cycle mutants

npj Systems Biology and Applications ◽

10.1038/s41540-020-0134-z ◽

2020 ◽

Vol 6 (1) ◽

Author(s):

Jenna E. Gallegos ◽

Neil R. Adames ◽

Mark F. Rogers ◽

Pavel Kraikivski ◽

Aubrey Ibele ◽

...

Keyword(s):

Cell Cycle ◽

Yeast Cell ◽

High Throughput ◽

Genetic Interactions ◽

Yeast Cell Cycle ◽

High Throughput Phenotyping

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Abundances of transcripts, proteins, and metabolites in the cell cycle of budding yeast reveal coordinate control of lipid metabolism

Molecular Biology of the Cell ◽

10.1091/mbc.e19-12-0708 ◽

2020 ◽

Vol 31 (10) ◽

pp. 1069-1084 ◽

Cited By ~ 3

Author(s):

Heidi M. Blank ◽

Ophelia Papoulas ◽

Nairita Maitra ◽

Riddhiman Garge ◽

Brian K. Kennedy ◽

...

Keyword(s):

Cell Cycle ◽

Lipid Metabolism ◽

Cell Division ◽

Yeast Cell ◽

Budding Yeast ◽

Yeast Cells ◽

Dynamic Changes ◽

Coordinate Control ◽

Cycle Dependent ◽

Budding Yeast Cell Cycle

In several systems, including budding yeast, cell cycle-dependent changes in the transcriptome are well studied. In contrast, few studies queried the proteome during cell division. There is also little information about dynamic changes in metabolites and lipids in the cell cycle. Here, the authors present such information for dividing yeast cells.

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A Role for Cell Polarity in Lifespan and Mitochondrial Quality Control in the Budding Yeast Saccharomyces cerevisiae

10.1101/2021.09.30.462607 ◽

2021 ◽

Author(s):

Emily J. Yang ◽

Wolfgang M Pernice ◽

Liza A. Pon

Keyword(s):

Cell Cycle ◽

Quality Control ◽

Cell Division ◽

Yeast Cell ◽

Cell Polarity ◽

Yeast Saccharomyces Cerevisiae ◽

A Cell ◽

Mitochondrial Distribution ◽

Mother Cells ◽

F Box Protein

SummaryBabies are born young, largely independent of the age of their mothers. Mother-daughter age asymmetry in yeast is achieved, in part, by inheritance of higher-functioning mitochondria by daughter cells and retention of some high-functioning mitochondria in mother cells. The mitochondrial F-box protein, Mfb1p, tethers mitochondria at both poles in a cell cycle-regulated manner: it localizes to and anchors mitochondria to the mother cell tip throughout the cell cycle, and to the bud tip prior to cytokinesis. Here, we report that cell polarity and polarized localization of Mfb1p decline with age in S. cerevisiae. Moreover, deletion of BUD1/RSR1, a Ras protein required for cytoskeletal polarization during asymmetric yeast cell division, results in depolarized Mfb1p localization, defects in mitochondrial distribution and quality control, and reduced replicative lifespan. Our results demonstrate a role for the polarity machinery in lifespan through modulating Mfb1 function in asymmetric inheritance of mitochondria during yeast cell division.

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Genetic Control of the Cell Division Cycle in Yeast: A model to account for the order of cell cycle events is deduced from the phenotypes of yeast mutants

Science ◽

10.1126/science.183.4120.46 ◽

1974 ◽

Vol 183 (4120) ◽

pp. 46-51 ◽

Cited By ~ 640

Author(s):

L. H. Hartwell ◽

J. Culotti ◽

J. R. Pringle ◽

B. J. Reid

Keyword(s):

Cell Cycle ◽

Cell Division ◽

Genetic Control ◽

Cell Division Cycle ◽

Yeast Mutants

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CRISPR Interference for Rapid Knockdown of Essential Cell Cycle Genes inLactobacillus plantarum

mSphere ◽

10.1128/msphere.00007-19 ◽

2019 ◽

Vol 4 (2) ◽

Cited By ~ 9

Author(s):

Ine Storaker Myrbråten ◽

Kamilla Wiull ◽

Zhian Salehian ◽

Leiv Sigve Håvarstein ◽

Daniel Straume ◽

...

Keyword(s):

Cell Cycle ◽

Cell Division ◽

Rna Binding ◽

Rna Binding Proteins ◽

Essential Genes ◽

Content Type ◽

Genetic Tools ◽

Guide Rna ◽

Cell Cycle Genes ◽

Key Species

ABSTRACTStudies of essential genes in bacteria are often hampered by the lack of accessible genetic tools. This is also the case forLactobacillus plantarum, a key species in food and health applications. Here, we develop a clustered regularly interspaced short palindromic repeat interference (CRISPRi) system for knockdown of gene expression inL. plantarum. The two-plasmid CRISPRi system, in which a nuclease-inactivated Cas9 (dCas9) and a gene-specific single guide RNA (sgRNA) are expressed on separate plasmids, allows efficient knockdown of expression of any gene of interest. We utilized the CRISPRi system to gain initial insights into the functions of key cell cycle genes inL. plantarum. As a proof of concept, we investigated the phenotypes resulting from knockdowns of the cell wall hydrolase-encodingacm2gene and of the DNA replication initiator genednaAand ofezrA, which encodes an early cell division protein. Furthermore, we studied the phenotypes of three cell division genes which have recently been functionally characterized in ovococcal bacteria but whose functions have not yet been investigated in rod-shaped bacteria. We show that the transmembrane CozE proteins do not seem to play any major role in cell division inL. plantarum. On the other hand, RNA-binding proteins KhpA and EloR are critical for proper cell elongation in this bacterium.IMPORTANCEL. plantarumis an important bacterium for applications in food and health. Deep insights into the biology and physiology of this species are therefore necessary for further strain optimization and exploitation; however, the functions of essential genes in the bacterium are mainly unknown due to the lack of accessible genetic tools. The CRISPRi system developed here is ideal to quickly screen for phenotypes of both essential and nonessential genes. Our initial insights into the function of some key cell cycle genes represent the first step toward understanding the cell cycle in this bacterium.

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Rv0954 Is a Member of the Mycobacterial Cell Division Complex

Frontiers in Microbiology ◽

10.3389/fmicb.2021.626461 ◽

2021 ◽

Vol 12 ◽

Author(s):

Ruojun Wang ◽

Sabine Ehrt

Keyword(s):

Cell Cycle ◽

Cell Wall ◽

Mycobacterium Tuberculosis ◽

Cell Division ◽

Cell Morphology ◽

Transposon Mutagenesis ◽

Genetic Interactions ◽

Intracellular Pathogen ◽

Resistance To Antibiotics ◽

Related Proteins

Proper control of cell division in the intracellular pathogen Mycobacterium tuberculosis is central to its growth, survival, pathogenesis, and resistance to antibiotics. Nevertheless, the divisome components and mechanisms by which mycobacteria regulate their cell cycle are not entirely understood. Here we demonstrate that the previously uncharacterized Rv0954 protein localizes to the mid-cell during cell division and interacts with the division-related proteins LamA, PbpA, and PknH. Deletion of rv0954 did not result in alterations in cell morphology or sensitivity to cell wall-targeting antibiotics but transposon mutagenesis demonstrated genetic interactions with genes related to cell division. This work suggests that Rv0954 participates in cell division and reveals potential components of the mycobacterial divisome for future investigation.

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MUTE Directly Orchestrates Cell State Switch and the Single Symmetric Division to Create Stomata

10.1101/286443 ◽

2018 ◽

Cited By ~ 1

Author(s):

Soon-Ki Han ◽

Xingyun Qi ◽

Kei Sugihara ◽

Jonathan H. Dang ◽

Takaho A. Endo ◽

...

Keyword(s):

Gene Expression ◽

Cell Cycle ◽

Cell Division ◽

List Type ◽

Transcriptional Repressors ◽

Potent Inducer ◽

Feed Forward ◽

Feed Forward Loop ◽

Symmetric Division ◽

Cell Cycle Genes

SUMMARYPrecise cell division control is critical for developmental patterning. For the differentiation of a functional stoma, a cellular valve for efficient gas exchange, the single symmetric division of an immediate precursor is absolutely essential. Yet, the mechanism governing the single division event remains unclear. Here we report the complete inventories of gene expression by the Arabidopsis bHLH protein MUTE, a potent inducer of stomatal differentiation. MUTE switches the gene expression program initiated by its sister bHLH, SPEECHLESS. MUTE directly induces a suite of cell-cycle genes, including CYCD5;1, and their transcriptional repressors, FAMA and FOUR LIPS. The architecture of the regulatory network initiated by MUTE represents an Incoherent Type 1 Feed-Forward Loop. Our mathematical modeling and experimental perturbations support a notion that MUTE orchestrates a transcriptional cascade leading to the tightly-restricted, robust pulse of cell-cycle gene expression, thereby ensuring the single cell division to create functional stomata.HighlightsComplete inventories of gene expression in stomatal differentiation state are elucidatedMUTE switches stomatal patterning program initiated by its sister bHLH, SPEECHLESSMUTE directly induces cell-cycle genes and their direct transcriptional repressorsIncoherent feed-forward loop by MUTE ensures the single division of a stomatal precursor

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Synergistic CDK control pathways maintain cell size homeostasis

10.1101/2020.11.25.397943 ◽

2020 ◽

Author(s):

James O. Patterson ◽

Souradeep Basu ◽

Paul Rees ◽

Paul Nurse

Keyword(s):

Cell Division ◽

Cell Size ◽

High Throughput ◽

Small Cells ◽

Control Network ◽

Size Information ◽

Division Cell ◽

Diploid Cells ◽

High Cyclin

AbstractTo coordinate cell size with cell division, cell size must be computed by the cyclin-CDK control network to trigger division appropriately. Here we dissect determinants of cyclin-CDK activity using a novel high-throughput single-cell in vivo system. We show that inhibitory phosphorylation of CDK encodes cell size information and works synergistically with PP2A to prevent division in smaller cells. However, even in the absence of all canonical regulators of cyclin-CDK, small cells with high cyclin-CDK levels are restricted from dividing. We find that diploid cells of equivalent size to haploid cells exhibit lower CDK activity in response to equal cyclin-CDK enzyme concentrations, suggesting that CDK activity is reduced by DNA concentration. Thus, multiple pathways directly regulate cyclin-CDK activity to maintain robust cell size homeostasis.

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