Masters Theses

Date of Award

12-2021

Degree Type

Thesis

Degree Name

Master of Science

Major

Biochemistry and Cellular and Molecular Biology

Major Professor

Gladys Alexandre

Committee Members

Elizabeth M. Fozo, Brad M. Binder, Maitreyi E. Das, Andreas Nebenfuehr

Abstract

Chemotaxis allows bacteria to navigate chemical gradients, providing a critical survival advantage in dynamic environments. Best defined in Escherichia coli, chemotaxis uses methyl-accepting chemotaxis receptors (chemoreceptors) to sense and relay sensory information to flagella. Chemoreceptor signaling arrays assemble as large clusters at the cell poles and are coupled to a phosphorylation signal transduction cascade that biases flagellar motor rotation, causing cells to tumble away from repellant stimuli. The E. coli model defines only a single chemotaxis pathway with five chemotaxis receptors. Most motile bacteria have multiple chemotaxis systems and many chemotaxis receptors, including the alphaproteobacterium Azospirillum brasilense. A. brasilense is an ubiquitous soil bacterium whose genome encodes 51 chemoreceptors and 4 chemotaxis pathways (Che1-4). Che1 and Che4 are chemotaxis systems that regulate swimming speed and reversal probability, respectively. Che2 and Che3 are uncharacterized chemotaxis-like (chemosensory) systems, predicted to regulate functions other than flagellar motility bias. Previous experimental evidence suggests that Che3 functions to control flocculation in A. brasilense, while Che2 is predicted to modulate flagellar biosynthesis based on homology to a closely related system in Rhodospirillum centenum.

In this work, mutagenesis, fluorescent microscopy, and behavioral assays were used to study the localization, potential interactions, and sensory function of the Tlp1, Tlp4a, Tlp73, Aer, and AerC A. brasilense chemoreceptors. We demonstrate the localization of Tlp1 and Tlp4a into polar arrays depends on the presence of the soluble AerC chemoreceptor, suggesting a structural role for AerC. We show that Aer does not localize to polar chemoreceptor clusters despite being previously shown to be essential for chemotaxis. We also provide evidence that the uncharacterized chemoreceptor, Tlp73, is essential for aerotaxis in A. brasilense. Finally, using mutagenesis and behavioral assays, we address the potential function of the Che2 system. We provide evidence Che2 modulates polar flagellum rotation, not flagellar biosynthesis. We also provide preliminary evidence for crosstalk between the Che2 and Che3 systems. Together, these data suggest a model of an interconnected network for the A. brasilense chemoreceptors, chemotaxis, and chemosensory signaling pathways.

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