|Title:||TetR-dependent gene regulation in intracellular Listeria monocytogenes demonstrates the spatiotemporal surface distribution of ActA|
|Authors :||Schmitter, Sibylle|
Loessner, Martin J.
|Published in :||Molecular microbiology|
|Publisher / Ed. Institution :||Wiley|
|License (according to publishing contract) :||Licence according to publishing contract|
|Type of review:||Peer review (Publication)|
|Subjects :||Actins; Animals; Bacterial proteins; Cell culture techniques; Cell movement; Cytoplasm; Dipodomys; Bacterial gene expression regulation; Listeria monocytogenes; Membrane proteins; Rats; Spatio-temporal analysis; Tetracycline resistance; Virulence factors|
|Subject (DDC) :||570: Biology|
|Abstract:||To enable specific and tightly controlled gene expression both in vitro and during the intracellular lifecycle of the pathogen Listeria monocytogenes, a TetR-dependent genetic induction system was developed. Highest concentration of cytoplasmic TetR and best repression of tetO-controlled genes was obtained by tetR expression from the synthetic promoter Pt17 . Anhydrotetracycline (ATc) as inducer permitted concentration-dependent, fine-tuned expression of genes under control of the tetO operator and a suitable promoter. The actin-polymerizing ActA protein represents a major virulence factor of L. monocytogenes, required for actin-based motility and cell-to-cell spread in infected host cells. To be able to observe its spatial and temporal distribution on intracellular L. monocytogenes cells, conditional mutants featuring actA placed under TetR control were used to infect PtK2 epithelial cells. Following induction at different time intervals, the subsequent recruitment of actin by L. monocytogenes could be monitored. We found that cells displayed functional ActA after approximately 15 min, while formation of polarized actin tail was complete after 90-120 min. At this point, intracellular motility of the induced mutants was indistinguishable from wild-type bacteria. Interestingly, de novo ActA synthesis in intracellular Listeria also demonstrated the temporal, asymmetric redistribution of the membrane-anchored proteins from the lateral walls toward the cell poles.|
|Departement:||Life Sciences and Facility Management|
|Organisational Unit:||Institute of Food and Beverage Innovation (ILGI)|
|Publication type:||Article in scientific Journal|
|Appears in Collections:||Publikationen Life Sciences und Facility Management|
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