Abstract
Bactofilins are rigid, non-polar bacterial cytoskeletal filaments that link cellular processes to specific curvatures of the cytoplasmic membrane. Although homologs of bactofilins have been identified in archaea and eukaryotes, functional studies have remained confined to bacterial systems. Here, we characterized representatives of two families of archaeal bactofilins from the pleomorphic archaeon Haloferax volcanii, halofilin A (HalA) and halofilin B (HalB). Unlike bacterial bactofilins, HalA polymerizes into polar filaments in vivo at positive membrane curvature, whereas HalB forms more static foci and accumulates in areas of local negative curvatures on the outer cell surface. Combining gene deletions, super-resolution, and single-cell microscopy showed that halofilins are critical in maintaining H. volcanii cell integrity during shape transition from disk (sessile) to rod (motile). Morphological defects in ΔhalA primarily affected rod-shaped cells from accumulating highly positive curvatures. Conversely, disk-shaped cells were exclusively affected by halB deletion, showing a decrease in positive and negative curvatures, resulting in flatter cells. Furthermore, while ΔhalA and ΔhalB cells displayed lower cell division placement precision, morphological defects arose predominantly during the disk-to-rod shape remodeling. We propose halofilins provide mechanical scaffolding, dynamically coupling the cytoplasmic membrane and the S-layer. We speculate that HalA filaments support rods under low S-layer lipidation (flexible, fast membrane diffusion). In contrast, HalB connects the S-layer to negative curvatures in disks under high lipidation levels (rigid, slow membrane diffusion).
Significance Statement How have cells evolved molecular machineries to support the mechanical and morphological requirements of microbial behavior? The biochemistry and biophysics of cell envelopes and their associated cytoskeleton are key to understanding such adaptations. Archaea are strategically positioned in the Tree of Life, bridging the molecular and cellular biology of bacteria and eukaryotes. Our work investigated the archaeal shapeshift between two morphological cell types and the role of two new cytoskeletal proteins in mechanical scaffolding during development. Specifically, halofilins are the first identified archaeal cell shape factors to be demonstrated to function at the onset of cell-type shifting. Broadly, our model offers potential for new synthetic cell designs and bioinspired materials that are both minimalist and modular.
Competing Interest Statement
The authors have declared no competing interest.
Footnotes
Author Contributions: AB, VA, MP, ZC, PE, and JM designed research; ZC, PE, JM, TR, OL, AD, KA, JK, and KY performed research; ZC, JM, OL, JK, KY and MP contributed new reagents/analytic tools; ZC, PE, JM, TR, OL, AD, and KY analyzed data; all authors contributed to the manuscript writing and revision.
Competing Interest Statement: The authors declare no competing interest.
Classification: Biological Sciences
Subgroup: Cell Biology