Complexity in Rhodobacter sphaeroides chemotaxis

Thesis

Perceiving and responding to the environment is key to survival. Using the prokaryotic equivalent of a nervous system – the chemotaxis system – bacteria sense chemical stimuli and respond by adjusting their movement accordingly.

In chemotactic bacteria, such as the well-studied E. coli, environmental nutrient sensing is achieved through a membrane embedded protein array that specifically clusters at the cell poles. Signalling to the motor is performed by activation of the CheA kinase, which phosphorylates CheY and CheB. CheY–P tunes the activity of the flagellar motor while CheB–P, together with CheR is involved in adaptation to the stimulus. In E. coli, a dedicated phosphatase terminates the signal.

Most bacterial species however, have a much more complex chemotaxis network. Rhodobacter sphaeroides, a model organism for complex chemotaxis systems, has one membrane-embedded chemosensory array and one cytoplasmic chemosensory array, plus several homologs of the E. coli chemotaxis proteins. Signals from both arrays are integrated to control the rotation of a single start-stop flagellar motor.

The phosphorelay network has been studied extensively through in vitro phosphotransfer while in vivo studies have established the components of each array and the requirements for formation. Mathematical modelling has also contributed towards inferring connectivities within the signalling network.

Starting by constructing a two-hybrid-based interaction network focused on the components of the cytoplasmic chemosensory array, this thesis further addresses its associated adaptation network through a series of in vivo techniques.

The swimming behaviour of series of deletion mutants involving the adaptation network of R. sphaeroides is characterised under steady state conditions as well as upon chemotactic stimulation. New connectivities within the R. sphaeroides chemotaxis network are inferred from analysing these data together with results from in vivo photoactivation localisation microscopy of CheB₂.

The experimental results are used to propose a new model for chemotaxis in R. sphaeroides.

Authors: Szollossi, A

• Type of award: DPhil
• Level of award: Doctoral
• Awarding institution: University of Oxford
• Language: English
• Keywords: PALM; rhodobacter sphaeroides; bacterial two-hybrid; free swimming; bacterial chemotaxis; complex chemotaxis; BACTH; chemotaxis; tethered cell
• Subjects: Systems Biology; Microbiology