Open Collections

Abstract

Endospore formation (sporulation) is an essential strategy for preserving genetic material under harsh environmental conditions employed by bacteria of the phylum Firmicutes. This process involves asymmetric cell division which produces two compartments, a mother cell and a dormant spore. The mature spore is surrounded by two lipid bilayers, the inner (IsM) and outer (OsM) spore membranes, derived from the cytoplasmic/inner membrane of the mother cell. In well-characterized monoderm species such as Bacillus subtilis, the spore discards the OsM upon germination, resulting in a vegetative cell surrounded by a single membrane. The recent discovery and characterization of diderm spore-formers, such as the Negativicute Acetonema longum, reveal an unprecedented retention of the OsM and its subsequent remodelling into a canonical outer membrane (OM), containing lipopolysaccharide (LPS) and β-barrel proteins (OMPs). In this study, we sought to elucidate the mechanisms of cell envelope remodelling in A. longum by analyzing the peptidoglycan (PG), LPS and OMP biosynthetic pathways throughout the sporulation cycle. Our study shows that, despite the overall typical diderm architecture, the A. longum cell envelope possesses non-canonical and possibly ancient components involved in membrane tethering, LPS transport and OMP translocation. High-resolution imaging, biochemical and omics approaches further reveal that OM biogenesis commences during germination and occurs de novo. Based on these findings, we propose a model for cell envelope remodelling in sporulating diderms, which involves stage-specific cell wall synthases and hydrolases, and a novel mechanism for OMP insertion. Because A. longum and other members of the class Negativicutes possess a typical OM and are able to sporulate, they may represent an evolutionary missing link between the monoderm and diderm bacteria. As such, our study helps narrow the knowledge gap between the monoderms and diderms, providing insight into the major evolutionary events that have shaped extant bacterial phyla.