Biology subjects

Meyer, F. M.

Publications and source records attributed to Meyer, F. M..

3 recordsLinked to original sources

bioRxiv2025

The ParA-like ATPase PldP influences the TatA dynamics in Corynebacterium glutamicum

In bacterial cells, precise localization of protein complexes is achieved by unique positioning systems. One of the examples of such positioning systems is the ParAB-parS which is responsible for plasmid and chromosome segregation. In Corynebacterium glutamicum, a parAB deletion results in cell division and growth defects, while deletion of an orphan ParA-like protein pldP results only in a moderate cell division phenotype. Having confirmed a basal ATPase activity of PldP, we aimed to explore if the {Delta}pldP-related phenotype could be a consequence of the mislocalized secreted proteins, as the loss of extracellular proteins involved in cell wall metabolism results in a similar phenotype characterized by disrupted separation of daughter cells. Putative peptidoglycan hydrolase Rv2525c from Mycobacterium tuberculosis and Rv2525c-like glycoside hydrolase-like domain-containing protein Cg0955 from C. glutamicum were previously shown to be transported outside of the cell by twin-arginine protein translocation machinery (Tat). Here, we found that although the deletion of pldP did not lead to the altered secretion of the putative hydrolase Cg0955 by the Tat system, it resulted in the reduction of the Tat dynamics. Our findings highlight the interplay between the ParA-like ATPase PldP and the Tat translocon and contribute to the studies of ParA-like proteins being essential in positioning various cargos in the bacterial cells. ImportancePrecise spatio-temporal localization of protein complexes within a bacterial cell is essential for the survival and proliferation of bacteria. ParA-like ATPases play a crucial role in protein positioning, as well as chromosome and plasmid segregation. Here, we characterize a novel ParA-like ATPase PldP in Corynebacterium glutamicum, a model organism for the cell biology of Mycobacteriales and a biotechnological workhorse. Deletion of pldP results in the cell division phenotypes and impacts the intracellular dynamics of TatA, a component of the twin-arginine protein transport. We suggest that the mislocalization of the Tat-secreted putative peptidoglycan hydrolase caused by the indirect influence of pldP deletion might account for the observed cell separation defect.

microbiology↗

bioRxiv2023

Deletion of SMC renders FtsK essential in Corynebacterium glutamicum

Structural maintenance of chromosomes (SMC) are ubiquitously distributed proteins involved in chromosome organization. Deletion of smc causes severe growth phenotypes in many organisms. Surprisingly, smc can be deleted in Corynebacterium glutamicum, a member of the Actinomycetota phylum, without any apparent growth phenotype. Earlier work has shown that SMC in C. glutamicum is loaded in a ParB-dependent fashion to the chromosome and functions in replichore cohesion. The unexpected absence of a growth phenotype in the smc mutant prompted us to screen for unknown synthetic interactions within C. glutamicum. Therefore, we generated a high-density Tn-5 library based on wild-type and smc-deleted C. glutamicum strains. The transposon sequencing (Tn-seq) data revealed that the DNA-translocase FtsK is essential in a smc deletion strain. FtsK localized to the septa and cell poles in wild type cells, however deletion of smc resulted in a decreased polar FtsK localization. Single-particle tracking analysis further suggests that prolonged FtsK complex activity is both required and sufficient to make up for the absence of SMC, thus achieving efficient chromosome segregation in C. glutamicum. Further, single molecule dynamics of FtsK is influenced, albeit indirectly, by DNA-loaded SMC. Deletion of ParB results in an increased of both SMC and FtsK mobility. While the first change agrees with previous data that show how ParB is essential for SMC loading on DNA, the latter suggests that FtsK mobility is affected in cells with defects in chromosome organization. Based on our data we propose a simple, yet efficient mechanism for efficient DNA segregation in C. glutamicum, even in absence of SMC proteins. ImportanceFaithful DNA segregation is of fundamental importance for life. Bacteria have efficient systems to coordinate chromosome compaction, DNA segregation and cell division. A key factor in DNA compaction is the SMC-complex that is found to be essential in many bacteria. In members of the Actinomycetota smc is dispensable, but the reason for this was unclear. We show here that the divisome associated DNA-pump FtsK can compensate SMC loss and the subsequent loss in correct chromosome organization. In cells with distorted chromosomes, FtsK functions for an extended period of time at the septum, until chromosomes are segregated.

microbiology↗

bioRxiv2023

Effects of benzothiazinone and ethambutol on the integrity of the corynebacterial cell envelope

The mycomembrane (MM) is a hydrophobic layer formed by mycolic acids covering the surface of Mycobacteria and related species. This group includes important pathogens such as Mycobacterium tuberculosis, Corynebacterium diphtheriae, but also the biotechnologically important strain Corynebacterium glutamicum. The MM contributes to the impermeability of the cell envelope and thereby also protects bacteria from antibiotics. This makes biosynthesis of the MM an attractive target for antibiotic intervention. The first line anti-tuberculosis drug ethambutol (EMB) interferes with the synthesis of the arabinogalactan (AG), which is a structural scaffold for covalently attached mycolic acids that form the inner leaflet of the MM. Similarly, the new drug candidate, benzothiazinone 043 (BTZ) affects the synthesis of the AG component of the cell wall. We previously showed that C. glutamicum cells treated with a sublethal concentration of EMB lose the integrity of the MM. In this study we examined the effects of a sublethal concentration of BTZ. Our work shows that 1 {micro}g ml-1 BTZ efficiently blocks the apical growth machinery and reduces cell proliferation, however the integrity of the MM is largely preserved and the effects of {beta}-lactam antibiotics are only additive, not synergistic. Transmission electron microscopy (TEM) analysis revealed a distinct middle layer in the septum of control cells considered to be the inner leaflet of the MM covalently attached to the AG. It functions as a greasy slide for the lateral flow of mycolic acids in the outer leaflet. This layer was not detectable in the septa of BTZ or EMB treated cells, which suggests that the greasy slide is impaired and the confluency of the MM is thereby reduced. In addition, we observed that EMB treated cells have a thicker and less electron dense peptidoglycan (PG) layer consistent with the report that EMB also inhibits the glutamate racemase MurI. We conclude that EMB and BTZ have distinct differences in their modes of action. While EMB and BTZ both effectively block elongation growth, BTZ also strongly reduces septal cell wall synthesis. This renders BTZ treated cells likely more tolerant to antibiotics that act on growing bacteria.

microbiology↗