Biology subjects

Tan, W. B.

Publications and source records attributed to Tan, W. B..

5 recordsLinked to original sources

bioRxiv2025

Polar localization of putative phospholipid transporters in Escherichia coli

The Gram-negative bacterial cell envelope comprises an outer membrane (OM) with an asymmetric arrangement of lipopolysaccharides (LPS) and phospholipids (PLs), protecting them from both physical and chemical threats. To build the OM, PLs must be transported across the cell envelope; this process has remained elusive until recently, where three collectively essential AsmA-superfamily proteins - YhdP, TamB, and YdbH - are proposed to function as anterograde PL transporters in Escherichia coli. Here, we identify the cell wall-binding protein DedD as a novel interacting partner of YhdP, and discover that all three AsmA-superfamily proteins are recruited to, and strongly enriched at the cell poles. Our observation raises the possibility that anterograde PL transport could be spatially restricted to the cell poles, and highlights the importance of understanding the spatial-temporal regulation of OM biogenesis in coordination with cell growth and division.

microbiology↗

bioRxiv2024

Primary role of the Tol-Pal complex in bacterial outer membrane lipid homeostasis

Gram-negative bacteria are defined by an outer membrane (OM) that contributes to envelope integrity and barrier function. Building this bilayer require proper assembly of lipopolysaccharides, proteins, and phospholipids, yet how the balance of these components is achieved is unclear. One system long known for ensuring OM stability is the Tol-Pal complex, which has been implicated in maintaining OM lipid homeostasis. However, assignment of Tol-Pal function has been challenging, owing to its septal localization and associated role(s) during division. Here, we uncouple the function of Tol-Pal in OM lipid homeostasis from its impact on cell division in Escherichia coli, by engineering a chimeric complex that loses septal enrichment. We demonstrate that this peripherally-localized Tol-Pal complex is fully capable of maintaining lipid balance in the OM, thus restoring OM integrity and barrier. Our work establishes the primary function of the Tol-Pal complex in OM lipid homeostasis, independent of its role during division.

microbiology↗

bioRxiv2021

Genetic interaction mapping highlights key roles of the Tol-Pal complex

The conserved Tol-Pal trans-envelope complex is important for outer membrane (OM) stability and cell division in Gram-negative bacteria. It has been proposed to mediate OM constriction during cell division via tethering to the cell wall. Yet, recent studies suggest that the complex has additional roles in OM lipid homeostasis and septal cell wall separation. How the Tol-Pal complex functions to facilitate these many processes is unclear. To gain insights into its role(s), we applied transposon insertion sequencing, and report here a detailed network of genetic interactions with the tol-pal locus in Escherichia coli. We found one positive and >20 negative strong interactions based on fitness. Disruption of genes responsible for osmoregulated periplasmic glucan biosynthesis restores fitness and OM barrier function, but not cell division defects, in tol-pal mutants. In contrast, deletions of genes involved in OM homeostasis and cell wall remodelling give rise to synthetic growth defects in strains lacking Tol-Pal, especially exacerbating OM barrier and/or cell division defects. Notably, the {Delta}tolA mutant having additional defects in OM protein assembly ({Delta}bamB) exhibited severe division phenotypes, even under conditions where the single mutants divide normally; this highlights the possibility for OM phenotypes to indirectly influence the cell division process. Overall, our work provides insights into the intricate nature of Tol-Pal function, and reinforces the model that this complex plays crucial roles in cell wall-OM tethering, cell wall remodelling, and in particular, OM homeostasis.

microbiology↗

bioRxiv2021

YhcB coordinates peptidoglycan and LPS biogenesis with phospholipid synthesis during Escherichia coli cell growth.

The cell envelope is essential for viability in all kingdoms of life. It retains enzymes and substrates within a confined space while providing a protective barrier to the external environment. Destabilising the envelope of bacterial pathogens is a common strategy employed by antimicrobial treatment. However, even in one of the most well studied organisms, Escherichia coli, there remain gaps in our understanding of how the synthesis of the successive layers of the cell envelope are coordinated during growth and cell division. Here, we used a whole genome phenotypic screen to identify mutants with a defective cell envelope. We report that loss of yhcB, a conserved gene of unknown function, results in loss of envelope stability, increased cell permeability and dysregulated control of cell size. Using whole genome transposon mutagenesis strategies we report the complete genetic interaction network of yhcB, revealing all genes with a synthetic negative and a synthetic positive relationship. These genes include those previously reported to have a role in cell envelope biogenesis. Surprisingly, we identified genes previously annotated as essential that became non-essential in a {Delta}yhcB background. Subsequent analyses suggest that YhcB sits at the junction of several envelope biosynthetic pathways coordinating the spatiotemporal growth of the cell, highlighting YhcB as an as yet unexplored antimicrobial target.

microbiology↗

bioRxiv2018

Biosynthetic intermediates of the enterobacterial common antigen overcome outer membrane lipid dyshomeostasis in Escherichia coli

The outer membrane (OM) is an essential component of the Gram-negative bacterial envelope that protects cells against external threats. To maintain a functional OM, cells require distinct mechanisms to ensure balance of proteins and lipids in the membrane. Mutations in OM biogenesis and/or homeostasis pathways often result in permeability defects, but how molecular changes in the OM affect barrier function is unclear. Here, we seek potential mechanism(s) that can alleviate permeability defects in Escherichia coli cells lacking the Tol-Pal complex, which accumulate excess PLs in the OM. We identify mutations in enterobacterial common antigen (ECA) biosynthesis that re-establish OM barrier function against large hydrophilic molecules, yet did not restore lipid homeostasis. Furthermore, we demonstrate that build-up of biosynthetic intermediates, but not loss of ECA itself, contributes to the rescue. This suppression of OM phenotypes is unrelated to known effects that accumulation of ECA intermediates have on the cell wall. Finally, we reveal that an unusual diacylglycerol pyrophosphoryl-linked lipid species also accumulates in ECA mutants, and might play a role in the rescue phenotype. Our work provides insights into how OM barrier function can be restored independent of lipid homeostasis, and highlights previously unappreciated effects of ECA-related species in OM biology.

microbiology↗