Curved bacteria represent one of the most common of bacterial shapes yet the mechanisms by which
bacteria become curved and the functions of curvature are largely unknown. In addition, large-scale polymers
assemble in both the cytoplasm and periplasm yet only the cytoplasmic polymers have been well characterized
to date. This is an important distinction, as periplasmic proteins must contend with different molecular
environments like oxidizing conditions and a lack of ATP/GTP energy sources. My lab recently demonstrated
that two proteins, CrvA and CrvB, assemble into periplasmic structures that are essential for establishing cell
curvature in the pathogen, Vibrio cholerae. Here we propose to leverage this system to address the field’s
gaps in understanding both curved shapes and periplasmic polymer formation.
In previous work, we identified and characterized the first curvature determinant in V. cholerae, CrvA.
CrvA forms polymers in the periplasm that pattern the insertion of new cell wall to cause these bacteria to
curve. More recently we discovered a second curvature determinant, CrvB. CrvA and CrvB colocalize in the V.
cholerae periplasm, and CrvA and CrvB co-expression are sufficient to induce curvature in straight Vibrio
species, E. coli, P. aeruginosa, and even distantly-related C. crescentus and A. tumefaciens.
Here we seek to better understand bacterial curvature determination and function by answering three
outstanding questions in three aims. 1) How do CrvA/B assemble in the periplasm, whose oxidizing conditions
and lack of ATP/GTP make it a very different environment from the cytoplasm where well-characterized
cytoskeletons assemble? Aim 1 will answer this question by combining electron and fluorescence microscopy
to determine the structure and dynamics of CrvA/CrvB assembly. 2) How do CrvAB actually generate cell
curvature? CrvAB are distinct from previously-characterized shape determinants in both being periplasmic and
functioning autonomously of other shape-patterning elements like MreB. Aim 2 will thus address how CrvA and
CrvB induce curvature by identifying and characterizing their interactions with the cell wall and other partners.
3) How does curvature affect bacterial behaviors? Aim 3 will harness our ability to synthetically curve bacteria
and use single-cell imaging to determine how curvature affects motility in gels, biofilms, and on surfaces.
These experiments will dissect the biophysical basis of important behaviors that result from cell curvature.
Together these studies will establish V. cholerae CrvAB as a powerful model system for studying cell
shape across multiple scales, answering fundamental questions to advance several fields. First, at the
molecular scale, we will learn how polymers form in the periplasm. Second, at the cellular scale, we will learn
how two proteins can curve an immense range of bacteria. And third, at the behavioral/evolutionary scale, we
will learn the benefits conferred by one of the most common of bacterial shapes.