Genome-wide phenotypic analysis of growth, cell morphogenesis and cell cycle events in Escherichia coli.

Campos, Manuel; Dobihal, Genevieve S; Jacobs‐Wagner, Christine

doi:10.1101/101832

Cited by 15 publications

(28 citation statements)

References 93 publications

(102 reference statements)

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“…intermediates is independent of und-P sequestration or σ E and Cpx stress response pathways Recently, it has been reported that the build-up of ECA "dead-end" intermediates can lead to sequestration of und-P, the common precursor for many sugar polymers in the cell envelope including peptidoglycan; this gives rise to shape defects such as filamentation and swelling Campos et al, 2018). In our strains that lack colanic acids, we also observe similar shape defects.…”

Section: Suppression Of Om Permeability Defects In Tol-pal Strains VIsupporting

confidence: 79%

Mutations in enterobacterial common antigen biosynthesis restore outer membrane barrier function inEscherichia coli tol-palmutants

Jiang

Tan

Shrivastava

et al. 2018

Preprint

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23The outer membrane (OM) is an essential component of the Gram-negative bacterial cell 24 envelope that protects cells against external threats such as antibiotics. To maintain a stable and 25 functional OM barrier, cells require distinct mechanisms to ensure a balance of proteins and 26 lipids in the membrane. Crucial to this is the proper transport and assembly of various OM 27 components, of which the process of phospholipid (PL) transport is least understood. How OM 28 assembly pathways are coordinated to achieve homeostasis is also unclear. In this study, we set 29 out to identify potential mechanism(s) that can alleviate OM lipid dyshomeostasis in Escherichia 30 coli. Cells lacking the Tol-Pal complex accumulate excess PLs in the OM due to defective 31 retrograde PL transport. Here, we isolated mutations in enterobacterial common antigen (ECA) 32 biosynthesis that restore OM barrier function in these strains; build-up of biosynthetic 33 intermediates along the ECA pathway is key to this rescue. Interestingly, these ECA mutations 34 re-establish OM lipid homeostasis in cells lacking the Tol-Pal complex yet do not act by 35 restoring retrograde PL transport. Furthermore, a novel diacylglycerol pyrophosphoryl-linked 36 ECA species structurally similar to PLs can be detected in the inner membrane of ECA mutants. 37 We therefore propose a model where these unique species may modulate anterograde PL 38 transport to overcome OM lipid dyshomeostasis. Our work provides insights into bacterial lipid 39 transport across the cell envelope and highlights previously unappreciated effects of ECA 40 intermediates in OM biology. 41 42 3 Author Summary 43 44Biological membranes define cellular boundaries, allow compartmentalization, and represent a 45 prerequisite for life; yet, our understanding of membrane biogenesis and stability remain 46 rudimentary. In Gram-negative bacteria, the outer membrane prevents entry of toxic substances, 47 conferring intrinsic resistance against many antibiotics. How the outer membrane is assembled, 48 specifically lipid trafficking processes are not well understood. How this membrane is stably 49 maintained is also unclear. In this study, we discovered that intermediates along the biosynthetic 50 pathway of an exopolysaccharide exhibit stabilizing effects on outer membranes with lipid 51 imbalance in Escherichia coli. Our work suggests that these intermediates modulate phospholipid 52 trafficking within the double-membrane cell envelope to achieve outer membrane lipid 53 homeostasis. Furthermore, it provides a starting point to begin identifying hitherto unknown 54 phospholipid transport systems in Gram-negative bacteria, which are potential targets for the 55 development of future antibiotics. 56 57 58 Gram-negative bacteria are surrounded by a multilayered cell envelope consisting of the 59 inner membrane (IM), the peptidoglycan layer, and the outer membrane (OM). This envelope 60 structure, in particular the OM, plays an essential role in preventing toxic molecules from 61 en...

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Section: Suppression Of Om Permeability Defects In Tol-pal Strains VIsupporting

confidence: 79%

Mutations in enterobacterial common antigen biosynthesis restore outer membrane barrier function inEscherichia coli tol-palmutants

Jiang

Tan

Shrivastava

et al. 2018

Preprint

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“…Thus, new methods (e.g., high-throughput microscopy, flow cytometry, etc.) have been used to measure morphology when screening or selecting for mutants (13)(14)(15)(16)(17)(18), resulting in the discovery of new factors connected to cell division and morphogenesis. However, many of the morphological phenotypes discovered in these reports have no mechanistic explanation, and some lack functional annotation altogether.…”

mentioning

confidence: 99%

YtfB, an OapA Domain-Containing Protein, Is a New Cell Division Protein in Escherichia coli

Jorgenson

Young

2018

J Bacteriol

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While screening the Pfam database for novel peptidoglycan (PG) binding modules, we identified the OapA domain, which is annotated to be a LysM-like domain. LysM domains bind PG and mediate localization to the septal ring. In the Gram-negative bacterium , an OapA domain is present in YtfB, an inner membrane protein of unknown function but whose overproduction causes cells to filament. Together, these observations suggested that YtfB directly affects cell division, most likely through its OapA domain. Here we show that YtfB accumulates at the septal ring and that its action requires the initiating division protein FtsZ and to a lesser extent ZipA, an early recruit to the septalsome. While the loss of YtfB had no discernable impact, a mutant lacking both YtfB and DedD (a known cell division protein) grew as filamentous cells. The YtfB OapA domain by itself also localized to sites of division, and this localization was enhanced by the presence of denuded PG glycans. Finally, the OapA domain bound PG, though binding did not depend on the formation of denuded glycans. Collectively, our findings demonstrate that YtfB is a cell division protein whose function is related to cell wall hydrolases. All living cells must divide in order to thrive. In bacteria, this involves the coordinated activities of a large number of proteins that work in concert to constrict the cell. Knowing which proteins contribute to this process and how they function is fundamental. Here we identify a new member of the cell division apparatus in the Gram-negative bacterium whose function is related to the generation of a transient cell wall structure. These findings deepen our understanding of bacterial cell division.

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“…Modest defects in cell division are well known to increase bacterial cell size—particularly length—without impacting growth rate [4,7,50,51,53], while defects in transcription severely curtail growth but have little detectable impact on morphology [6]. Most importantly, an exhaustive phenotypic analysis of the entirety of non-essential genes in E. coli by the Jacobs-Wagner group, failed to identify any evidence supporting a direct connection between growth rate and size [54].…”

Section: Discussionmentioning

confidence: 99%

Comprehensive analysis of central carbon metabolism reveals multiple connections between nutrients, biosynthetic capacity, and cell morphology inEscherichia coli

Pa²

2017

Preprint

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Bacterial morphology is a complex trait that is highly sensitive to changes in the environment. For heterotrophic organisms, such as Escherichia coli, increases in nutrient levels are frequently accompanied by several-fold increases in both size and growth rate. Despite the dramatic nature of these changes, how alterations in nutrient availability translate into changes in growth and morphology remains a largely open question. To understand the signaling networks coupling nutrient availability with size and shape, we examined the impact of deletions in the entirety of non-essential central carbon metabolic genes on E. coli growth rate and cell size. Our data reveal the presence of multiple metabolic nodes that play important yet distinctive roles in shaping the cell. Consistent with recent work from our lab and others, although both are sensitive to nutrient availability, size and growth rate vary independently. Cell width and length also appear to be independent phenomena, influenced by different aspects of central carbon metabolism. These findings highlight the diversity of factors that can impact cell morphology and provide a foundation for further studies.Author summaryOften taken for granted, the shape of bacterial cells is a complex trait that is highly sensitive to environmental perturbations. Nutrients in particular, strongly impact bacterial morphology together with growth rate. The ubiquitous, rod-shaped bacteria Escherichia coli increases both length and width several fold upon a shift from nutrient poor to nutrient rich medium, a change accompanied by an equally dramatic increase in growth rate. Central carbon metabolism is an obvious site for the integration of nutrient dependent signals that dictate cell size and shape. To develop a clearer picture of the molecular mechanisms coupling nutrient assimilation with cell growth and morphology, we screened the entirety of nonessential carbon metabolic genes for their contribution to growth rate and cell shape. Our data reveal the presence of multiple regulatory circuits coordinating different metabolic pathways with specific aspects of cell growth and morphology. Together, these data firmly establish a role for central carbon metabolism as an environmentally sensitive sculptor of bacterial cells.

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Genome-wide phenotypic analysis of growth, cell morphogenesis and cell cycle events in Escherichia coli.

Cited by 15 publications

References 93 publications

Mutations in enterobacterial common antigen biosynthesis restore outer membrane barrier function inEscherichia coli tol-palmutants

Mutations in enterobacterial common antigen biosynthesis restore outer membrane barrier function inEscherichia coli tol-palmutants

YtfB, an OapA Domain-Containing Protein, Is a New Cell Division Protein in Escherichia coli

Comprehensive analysis of central carbon metabolism reveals multiple connections between nutrients, biosynthetic capacity, and cell morphology inEscherichia coli

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