S-layer expansion is mediated by hoop-like cracks in the paracrystalline lattice during Bacillus anthracis growth

Surface layers (S-layers) are paracrystalline protein monolayers that envelop archaea and some bacteria, including Bacillus anthracis. S-layer subunits in B. anthracis are tethered to the underlying cell wall through glycopolymers that are covalently attached to the peptidoglycan (PG) meshwork. Cell growth in this rod-shaped organism occurs by dispersed PG synthesis throughout the cell cylinder. How the crystalline lattice expands in lockstep with the cell wall has remained unclear. Here, using fluorescently labelled S-layer subunits and high-resolution imaging, we show that S-layer expansion during B. anthracis growth is mediated by pervasive hoop-like cracks in the lattice. Pulse-chase experiments reveal two modes of S-layer assembly at these cracks: dispersed and focused. Our data suggest that local pulling forces on the S-layer array generated by dispersed cell wall synthesis and expansion of the underlying PG cause crack formation, with ensuing dispersed and focused synthesis enabling S-layer expansion with bacterial growth. Hoop-like cracking of the S-layer, filled either completely or partially, respectively leading to dispersed or focused modes of S-layer synthesis, enables S-layer expansion during Bacillus anthracis growth.