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Insights on the Control of Yeast Single-Cell Growth Variability by Members of the Trehalose Phosphate Synthase (TPS) Complex.

ABSTRACT: Single-cell variability of growth is a biological phenomenon that has attracted growing interest in recent years. Important progress has been made in the knowledge of the origin of cell-to-cell heterogeneity of growth, especially in microbial cells. To better understand the origins of such heterogeneity at the single-cell level, we developed a new methodological pipeline that coupled cytometry-based cell sorting with automatized microscopy and image analysis to score the growth rate of thousands of single cells. This allowed investigating the influence of the initial amount of proteins of interest on the subsequent growth of the microcolony. As a preliminary step to validate this experimental setup, we referred to previous findings in yeast where the expression level of Tsl1, a member of the Trehalose Phosphate Synthase (TPS) complex, negatively correlated with cell division rate. We unfortunately could not find any influence of the initial TSL1 expression level on the growth rate of the microcolonies. We also analyzed the effect of the natural variations of trehalose-6-phosphate synthase (TPS1) expression on cell-to-cell growth heterogeneity, but we did not find any correlation. However, due to the already known altered growth of the tps1? mutants, we tested this strain at the single-cell level on a permissive carbon source. This mutant showed an outstanding lack of reproducibility of growth rate distributions as compared to the wild-type strain, with variable proportions of non-growing cells between cultivations and more heterogeneous microcolonies in terms of individual growth rates. Interestingly, this variable behavior at the single-cell level was reminiscent to the high variability that is also stochastically suffered at the population level when cultivating this tps1? strain, even when using controlled bioreactors.

SUBMITTER: Arabaciyan S

PROVIDER: S-EPMC7876269 | biostudies-literature | 2021

REPOSITORIES: biostudies-literature

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Insights on the Control of Yeast Single-Cell Growth Variability by Members of the Trehalose Phosphate Synthase (TPS) Complex.

Arabaciyan Sevan S Saint-Antoine Michael M Maugis-Rabusseau Cathy C François Jean-Marie JM Singh Abhyudai A Parrou Jean-Luc JL Capp Jean-Pascal JP

Frontiers in cell and developmental biology 20210128

Single-cell variability of growth is a biological phenomenon that has attracted growing interest in recent years. Important progress has been made in the knowledge of the origin of cell-to-cell heterogeneity of growth, especially in microbial cells. To better understand the origins of such heterogeneity at the single-cell level, we developed a new methodological pipeline that coupled cytometry-based cell sorting with automatized microscopy and image analysis to score the growth rate of thousands ...[more]

PMID: 33585476

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Project description:The crenarchaeon Sulfolobus acidocaldarius has been described to synthesize trehalose via the maltooligosyltrehalose synthase (TreY) and maltooligosyltrehalose trehalohydrolase (TreZ) pathway, and the trehalose glycosyltransferring synthase (TreT) pathway has been predicted. Deletion mutant analysis of strains with single and double deletions of ΔtreY and ΔtreT in S. acidocaldarius revealed that in addition to these two pathways, a third, novel trehalose biosynthesis pathway is operative in vivo: the trehalose-6-phosphate (T6P) synthase/T6P phosphatase (TPS/TPP) pathway. In contrast to known TPS proteins, which belong to the GT20 family, the S. acidocaldarius TPS belongs to the GT4 family, establishing a new function within this group of enzymes. This novel GT4-like TPS was found to be present mainly in the Sulfolobales The ΔtreY ΔtreT Δtps triple mutant of S. acidocaldarius, which lacks the ability to synthesize trehalose, showed no altered phenotype under standard conditions or heat stress but was unable to grow under salt stress. Accordingly, in the wild-type strain, a significant increase of intracellular trehalose formation was observed under salt stress. Quantitative real-time PCR showed a salt stress-mediated induction of all three trehalose-synthesizing pathways. This demonstrates that in Archaea, trehalose plays an essential role for growth under high-salt conditions.IMPORTANCE The metabolism and function of trehalose as a compatible solute in Archaea was not well understood. This combined genetic and enzymatic approach at the interface of microbiology, physiology, and microbial ecology gives important insights into survival under stress, adaptation to extreme environments, and the role of compatible solutes in Archaea Here, we unraveled the complexity of trehalose metabolism, and we present a comprehensive study on trehalose function in stress response in S. acidocaldarius This sheds light on the general microbiology and the fascinating metabolic repertoire of Archaea, involving many novel biocatalysts, such as glycosyltransferases, with great potential in biotechnology.

Project description:BackgroundTrehalose (a-D-glucopyranosyl a-D-glucopyranoside) is a nonreducing disaccharide and is widely distributed in bacteria, fungi, algae, plants and invertebrates. In the study, the identification of trehalose-6-phosphate synthase (TPS) genes stress-related in cotton, and the genetic structure analysis and molecular evolution analysis of TPSs were conducted with bioinformatics methods, which could lay a foundation for further research of TPS functions in cotton.ResultsThe genome information of Gossypium raimondii (group D), G. arboreum L. (group A), and G. hirsutum L. (group AD) was used in the study. Fifty-three TPSs were identified comprising 15 genes in group D, 14 in group A, and 24 in group AD. Bioinformatics methods were used to analyze the genetic structure and molecular evolution of TPSs. Real-time PCR analysis was performed to investigate the expression patterns of gene family members. All TPS family members in cotton can be divided into two subfamilies: Class I and Class II. The similarity of the TPS sequence is high within the same species and close within their family relatives. The genetic structures of two TPS subfamily members are different, with more introns and a more complicated gene structure in Class I. There is a TPS domain(Glyco transf_20) at the N-terminal in all TPS family members and a TPP domain(Trehalose_PPase) at the C-terminal in all except GrTPS6, GhTPS4, and GhTPS9. All Class II members contain a UDP-forming domain. The responses to environmental stresses showed that stresses could induce the expression of TPSs but the expression patterns vary with different stresses.ConclusionsThe distribution of TPSs varies with different species but is relatively uniform on chromosomes. Genetic structure varies with different gene members, and expression levels vary with different stresses and exhibit tissue specificity. The upregulated genes in upland cotton TM-1 is significantly more than that in G. raimondii and G. arboreum L. Shixiya 1.

Dataset Information

Insights on the Control of Yeast Single-Cell Growth Variability by Members of the Trehalose Phosphate Synthase (TPS) Complex.

Publications

Insights on the Control of Yeast Single-Cell Growth Variability by Members of the Trehalose Phosphate Synthase (TPS) Complex.

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