Research
Cell Shape Control:
How do cells control their shape, using nanoscale proteins? What proteins are important and what do these proteins do? In E. coli we know many of the proteins necessary for rod shape. However, the functions of these proteins are unknown. Because these proteins
are often essential it is difficult to determine their function. Using genetics and microscopy we aim to determine the function of many of these essential cell shape genes.
To this end we are exploring the function of MreB through the use of an alanine-scanning mutagenesis library. This is being used to quantitatively determine the cell shape of different mutants through microscopy, the effects of growth, and resistance to A22, an MreB depolymeizing drug.
We are also probabing the function of MreD through the use of a suppressor screen to find rod shaped MreD mutants.
The Cytoskeleton and Physiology
Antibiotic Resistance:
Can cells bipass the need for MreB? We have found that there is a specific cell density that once cells reach they are able to grow at a normal rate, even in the presence of A22. When cells are treated earlier than this density the become much larger and develop vacuoles. We have further shown that mecillinam also displays a density dependency. Interestingly, A22 and mecillinam target to proteins, MreB and PBP2, that are involved in cell wall elongation.
Using a screen for mutants that are more sensitive to MreB we have found a link between cell division and elongation. Deletion of EnvC, the activator of cell wall hydrolases required to seperate daughter cells after division, is more sensitive to A22. During A22 treatment EnvC accumulates into bright foci suggesting a direct interaction between MreB and EnvC.
Metabolism:
A screen for mutants that are resistant to the MreB depolymerizing drug, A22, revealed that many genes involved in central carbon metabolism result in resistant cells. We hypothesize that when grown on amino acids (LB medium), increases in gluconeogenesis leads to resistance to MreB distruption by increasing the concentration of cell wall precursors. Metabalomics is currently being performed to determine the changes in cell wall intermediates in specific metabolic mutants compared to WT cells with and without A22. In review
Motilty:
We have found that if you distrupt the MreB cytoskeleton chemotaxis is inhibited. This project has two main goals. The first is to determine what role MreB is playing in chemotaxis. The second goal is to seperate the role of MreB in cell shape control from its role in chemotaxis. We plan to do this by screening a mutant MreB library for both cell shape effects and motilty defects.