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Synaptopodin is necessary for Shigella flexneri intercellular spread.

ABSTRACT: For many intracellular pathogens, their virulence depends on an ability to spread between cells of an epithelial layer. For intercellular spread to occur, these pathogens deform the plasma membrane into a protrusion structure that is engulfed by the neighboring cell. Although the polymerization of actin is essential for spread, how these pathogens manipulate the actin cytoskeleton in a manner that enables protrusion formation is still incompletely understood. Here, we identify the mammalian actin binding protein synaptopodin as required for efficient intercellular spread. Using a model cytosolic pathogen, Shigella flexneri, we show that synaptopodin increases actin recruitment around bacteria and stabilizes the position of the actin tail at the posterior pole of the bacteria. We show that synaptopodin presence enables protrusions to form and to resolve at a greater rate, indicating that the proper alignment of the tail enables the bacteria to push against the membrane with greater force. We demonstrate that synaptopodin recruitment around bacteria requires the bacterial protein IcsA, and we show that this recruitment is further enhanced in a type 3 secretion system dependent manner. Myosin X, known to enhance protrusion formation, is associated with more intracellular bacteria in the presence of synaptopodin. These data establish synaptopodin as required for intracellular bacteria to reprogram the actin cytoskeleton in a manner that enables efficient protrusion formation and enhance our understanding of the cellular function of synaptopodin.

Significance

Intercellular spread is essential for many cytosolic dwelling pathogens during their infectious life cycle. Despite knowing the steps required for intercellular spread, relatively little is known about the host-pathogen interactions that enable these steps to occur. Here, we identify a requirement for the actin binding protein synaptopodin during intercellular spread by cytosolic bacteria. We show synaptopodin is necessary for the organization and abundance of actin around bacteria. We also demonstrate synaptopodin is necessary for the formation of plasma membrane structures known as protrusions that are necessary for the movement of these bacteria between cells. Thus, these findings implicate synaptopodin as an important actin-binding protein for the virulence of intracellular pathogens that require the actin cytoskeleton and spread between cells.

SUBMITTER: Vickery JM

PROVIDER: S-EPMC10168286 | biostudies-literature | 2023 Apr

REPOSITORIES: biostudies-literature

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Synaptopodin is necessary for <i>Shigella flexneri</i> intercellular spread.

Vickery Jenna M JM Toperzer Jody D JD Raab Julie E JE Lenz Laurel L LL Colgan Sean P SP Russo Brian C BC

bioRxiv : the preprint server for biology 20230626

For many intracellular pathogens, their virulence depends on an ability to spread between cells of an epithelial layer. For intercellular spread to occur, these pathogens deform the plasma membrane into a protrusion structure that is engulfed by the neighboring cell. Although the polymerization of actin is essential for spread, how these pathogens manipulate the actin cytoskeleton in a manner that enables protrusion formation is still incompletely understood. Here, we identify the mammalian acti ...[more]

PMID: 37163027

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Project description:The intracellular human pathogen Shigella flexneri invades the colon epithelium, replicates to high cell density within the host cell, and then spreads to adjacent epithelial cells. When S. flexneri gains access to the host cytosol, the bacteria metabolize host cytosolic carbon using glycolysis and mixed acid fermentation, producing formate as a by-product. We show that S. flexneri infection results in the accumulation of formate within the host cell. Loss of pyruvate formate lyase (PFL; ?pflB), which converts pyruvate to acetyl coenzyme A (CoA) and formate, eliminates S. flexneri formate production and reduces the ability of S. flexneri to form plaques in epithelial cell monolayers. This defect in PFL does not decrease the intracellular growth rate of S. flexneri; rather, it affects cell-to-cell spread. The S. flexneri ?pflB mutant plaque defect is complemented by supplying exogenous formate; conversely, deletion of the S. flexneri formate dehydrogenase gene fdnG increases host cell formate accumulation and S. flexneri plaque size. Furthermore, exogenous formate increases plaque size of the wild-type (WT) S. flexneri strain and promotes S. flexneri cell-to-cell spread. We also demonstrate that formate increases the expression of S. flexneri virulence genes icsA and ipaJ Intracellular S. flexneriicsA and ipaJ expression is dependent on the presence of formate, and ipaJ expression correlates with S. flexneri intracellular density during infection. Finally, consistent with elevated ipaJ, we show that formate alters S. flexneri-infected host interferon- and tumor necrosis factor (TNF)-stimulated gene expression. We propose that Shigella-derived formate is an intracellular signal that modulates virulence in response to bacterial metabolism.IMPORTANCEShigella is an intracellular pathogen that invades the human host cell cytosol and exploits intracellular nutrients for growth, enabling the bacterium to create its own metabolic niche. For Shigella to effectively invade and replicate within the host cytoplasm, it must sense and adapt to changing environmental conditions; however, the mechanisms and signals sensed by S. flexneri are largely unknown. We have found that the secreted Shigella metabolism by-product formate regulates Shigella intracellular virulence gene expression and its ability to spread among epithelial cells. We propose that Shigella senses formate accumulation in the host cytosol as a way to determine intracellular Shigella density and regulate secreted virulence factors accordingly, enabling spatiotemporal regulation of effectors important for dampening the host immune response.

Dataset Information

Synaptopodin is necessary for Shigella flexneri intercellular spread.

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Publications

Synaptopodin is necessary for <i>Shigella flexneri</i> intercellular spread.

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Dataset Information

Synaptopodin is necessary for Shigella flexneri intercellular spread.

Significance

Publications

Synaptopodin is necessary for &lt;i&gt;Shigella flexneri&lt;/i&gt; intercellular spread.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

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Synaptopodin is necessary for <i>Shigella flexneri</i> intercellular spread.