Biology subjects

Manger, S.

Publications and source records attributed to Manger, S..

5 recordsLinked to original sources

bioRxiv2026

Cell-envelope conduits enable transfer of megabase-sized double-stranded DNA between cells of the nosocomial pathogen Acinetobacter baumannii

Horizontal gene transfer in Acinetobacter baumannii has been attributed to conjugative pili, transduction, and natural transformation. Here, we describe a previously unrecognized type of chromosomal DNA exchange mediated by cell-envelope conduits that establish direct cytoplasmic continuity between neighboring A. baumannii cells. Cryo-electron tomography reveals that these conduits, which were found in multiple clinical isolates, comprise an outer membrane, a peptidoglycan layer, and an inner membrane. The ~65 nm-thick conduits contain two ~2 nm-thick filaments, which super-resolution microscopy identifies as DNA. Interstrain horizontal gene transfer assays coupled with whole-genome sequencing demonstrate the transfer and homologous recombination of chromosomal segments up to 1.1 Mbp, corresponding to as much as 27% of the A. baumannii genome. Together, these findings provide direct structural and functional evidence for a conduit-mediated large-scale chromosomal DNA exchange, thus expanding the known repertoire of horizontal gene transfer strategies that may contribute to the remarkable genomic plasticity of A. baumannii.

microbiology↗

bioRxiv2025

In situ structure of a gap junction - stomatin complex

Gap junctions (GJ) are intercellular channels that mediate electrical signals and the transfer of small molecules. GJs are crucial for the functions of the brain, heart and other organs. While structures of purified homomeric GJs are available, we lack in situ structures. In vivo, GJs can form heteromers with different functionalities, and may associate with other proteins. Here, we analyzed Caenorhabditis elegans GJs by cryo-electron tomography and sub-tomogram averaging. We observed hexagonal arrays of GJs at cellular junctions in primary embryonal cell culture that displayed distinct wide and narrow conformations. Moreover, in about 20% of the observed channels, we found a cap-like, cytosolic protein assembly enclosing the channel pore. We propose that the cap-structure is formed by the stomatin UNC-1, which is known to interact with C. elegans GJs, and strengthen this hypothesis by matching AlphaFold3 models of UNC-1 multimers with our GJ average. Furthermore, expressing UNC-1 and the C. elegans innexin UNC-9 in HEK cells resulted in similar structures at cell-cell contacts. UNC-1/stomatin ring assemblies may affect GJ formation or functions like rectification, that might be evolutionarily conserved. Significance StatementGap junction (GJ) channels connect neighboring cells. Structures of (purified) GJs have been studied in vitro, but not in situ. We identified GJ channels in primary Caenorhabditis elegans cells by cryo-electron tomography, and analyzed their structure by sub-tomogram averaging. The channels transverse the membranes of connected cells, and AlphaFold3 (AF3) models of the GJ subunit UNC-9, assuming dodecamers, fit the experimentally obtained surface map well. We observed a cytosolic cap structure on the GJ channels. The stomatin protein UNC-1 is known to physically interact with UNC-9 GJs. AF3 models of UNC-1 hexadecamers fit the cap structure, indicating that it may be formed by UNC-1, providing a first idea how UNC-1 interacts with, and may functionally influence, GJ channels.

neuroscience↗

bioRxiv2024

Structural characterization of an essential lipoprotein of Mycoplasma pneumoniae

Mycoplasma pneumoniae is a human pathogen causing atypical community-acquired pneumonia. It serves as a model for a minimal cell and is notable for its absent cell wall, minimal genome, and use of antigenic variation to evade the host immune response. Here, we report the structures of the essential surface proteins Mpn444 and Mpn436 at 3.74 [A] and 3.65 [A] resolution, and the molecular architecture of the Mpn444 homotrimeric complex. We show that both proteins include a peptidyl-prolyl isomerase (PPIase) domain and a chaperone-like domain. In vitro PPIase activity assays suggest that Mpn444 and Mpn436 function as extracellular foldases in Mycoplasma species. Furthermore, both proteins are conserved across multiple Mycoplasma species. We built a composite model integrating previously reported interactions from crosslinking and cryo-ET data and we conclude that Mpn444 is responsible for the extracellular folding of nascent protein chains. Our work underscores the potential of Mpn444 and Mpn436 as a target for the development of novel strategies to treat mycoplasma infections. Author SummaryIn this study, we discovered that the essential proteins Mpn444 and Mpn436 of Mycoplasma pneumoniae act as extracellular PPIases. We solved the structures of Mpn444, Mpn436, and Mpn444s homotrimer by single-particle cryo-EM analysis, marking them as the first experimentally resolved lipoproteins from Mycoplasma pneumoniae. The structures contain a peptidyl-prolyl isomerase (PPIase) domain as well as a chaperone-like domain, and the proteins show PPIase activity and bind unfolded proteins in vitro. By combining our structural data with previously published cryo-electron tomography and crosslinking data from M. pneumoniae cells, we suggest that Mpn444, and potentially Mpn436, works in concert with the Sec translocon and the membrane-bound ribosome. This finding is particularly important as the extracellular folding of bacterial proteins has been a significant gap in our understanding.

biophysics↗

bioRxiv2023

P116 from Mycoplasma is a self-sufficient lipid uptake and delivery machinery

Lipid acquisition and transport are fundamental processes in all organisms. Here, we investigate the lipid uptake and delivery mechanism of the minimal model organism Mycoplasma pneumoniae. We show that the essential protein P116 can transport lipids between liposomes independently and without ATP consumption. Our structural data and molecular dynamics simulations reveal the mechanism by which the N-terminal region of P116 perturbs the membrane, the lipid transfer route, and the regulation of membrane binding by the cargo mass within P116s large hydrophobic cavity. When adequately filled with cargo, P116 undergoes a rapid conformational change that modulates membrane binding. Taken together, our results show that Mycoplasma developed one integrated lipid uptake and delivery machinery that simplifies the complex multi-protein pathways used by higher developed organisms.

molecular biology↗

bioRxiv2022

The immunodominant protein P116 is a passive transporter of cholesterol and other essential lipids

Many human pathogens need to extract lipids from their environment for survival and proliferation. How this is accomplished on a molecular level is largely unknown1. Here, we report a comprehensive structural and functional analysis of the previously uncharacterized protein P116 (MPN_213) from Mycoplasma pneumoniae, a human pathogen responsible for approximately 30% of community-acquired human pneumonia2. Single-particle cryo-electron microscopy of P116 at 3.3 [A] resolution reveals a homodimer with a core domain presenting a previously unseen fold. This fold creates a large cavity of [~]18,000 [A]3 with a hydrophobic internal surface that is accessible to solvent. Within the cavity ligands with a length of 10-19 [A] and a width of 4 [A] could be observed. These ligands were identified as the essential lipids phosphatidylcholine, sphingomyelin and cholesterol using mass spectrometry. When the cavity is emptied, the protein undergoes an extensive conformational change that can no longer accommodate lipids. When emptied P116 is incubated with high-density lipoproteins (HDLs) a net transfer of cholesterol is demonstrated by a radioactivity experiment and cryo-electron microscopy resolves a complex between P116 and HDL. Taken together, our results reveal the mechanism by which P116 extracts essential lipids from the host environment and possibly then delivers them into the membrane by a wringing movement. This mechanism may be precedential for other cholesterol-auxotrophic bacteria.

microbiology↗