The Yersinia pestis YscF needle plays a role in the regulation of type III secretion and translocation

Date of Award




Degree Name

Doctor of Philosophy (Ph.D.)


Microbiology and Immunology

First Committee Member

Gregory V. Plano, Committee Chair


Activation of bacterial virulence-associated type III secretion systems (T3SSs) requires direct contact between a bacterium and a eukaryotic cell. In Y. pestis, a cytosolic YopN-TyeA complex functions to block type III secretion (T3S) in the presence of extracellular calcium and prior to contact with a eukaryotic cell. The mechanism by which the bacterium senses extracellular calcium and/or cell contact and transmits these signals to the cytosolic YopN-TyeA complex is unknown. I report here that YscF, a small protein that polymerizes to form the external needle of the T3SS, is essential for the cell contact- and calcium-dependent regulation of T3S. Alanine-scanning mutagenesis was used to identify YscF mutants that secrete virulence proteins in the presence and absence of calcium and prior to contact with a eukaryotic cell. Y. pestis YscF mutants with amino acid replacements I13A, D28A and D46A resulted in constitutive (unregulated) secretion that required 2.5, 5 and 7.5 mM Ca2+, respectively, to block secretion. Interestingly, a YscF D28A D46A double mutant that exhibited constitutive T3S that could not be blocked by any amount of Ca2+ was unable to translocate secreted Yops across a eukaryotic membrane. These data indicate that the YscF needle is a multifunctional structure that participates in virulence protein secretion, in translocation of virulence proteins across eukaryotic membranes and in the cell contact- and calcium-dependent regulation of T3S.


Biology, Microbiology

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