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Characterization of factors mediating localization of the Shigella actin assembly protein IcsA to the bacterial pole

Posted on:2010-02-10Degree:Ph.DType:Dissertation
University:Harvard UniversityCandidate:Fixen, Kathryn RFull Text:PDF
GTID:1440390002987936Subject:Biology
Abstract/Summary:
Spatial organization within bacteria is fundamental to many cellular processes, including virulence, DNA replication, chromosome segregation, cell division, protein secretion, chemotaxis, gene transfer, adhesion, and motility. In bacterial bacilli, the bacterial pole is a site to which proteins that specialize in each of these functions localize. However, detailed molecular understanding of these pathways is incomplete, and the breadth of the mechanisms that are operative in polarity are undefined. Autotransporter proteins, which represent the largest group of secreted proteins in gram-negative bacteria, are outer membrane proteins that commonly play a role in virulence and are generally secreted at the bacterial pole. The Shigella autotransporter IcsA localizes to the bacterial old pole where it mediates assembly of a propulsive actin tail inside infected mammalian host cells. Localization to the pole occurs in the cytoplasm prior to secretion. Here I helped develop a genetic reporter assay for protein localization to the pole, and using it I demonstrated that FtsQ possibly in complex with FtsB and FtsL, which are cell division proteins of unknown function, but not cell division per se, is required for localization of IcsA to the pole. The mechanism that establishes polar positional information in the cytoplasm depends on an extracytoplasmic activity via the periplasmic domain of FtsQ. A second autotransporter, SepA, also requires FtsQ for polar localization, indicating that FtsQ and its extracytoplasmic activity may have a common role in polar targeting of autotransporters.;I also used the genetic reporter assay to carry out mutagenic analysis of IcsA to determine residues required for its localization to the pole. A number of residues may be required for polar localization, but further analysis will be necessary to confirm these findings. In addition, these mutations could be used to find compensatory extragenic mutations that restore polar localization of IcsA. These mutations will likely be in genes encoding proteins that interact with IcsA, which will help further characterize this conserved mechanism of polar localization.
Keywords/Search Tags:Localization, Icsa, Protein, Pole, Bacterial, Cell division
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