Biology subjects

Rosenshine, I.

Publications and source records attributed to Rosenshine, I..

6 recordsLinked to original sources

bioRxiv2026

A small RNA guides a post-transcriptional regulatory protein to its target

RNA-binding proteins are central to post-transcriptional regulation and are generally thought to recognize their target RNAs through direct binding. Here, we uncover a previously unrecognized mechanism in which a small RNA (sRNA) guides a protein regulator to specific target mRNAs, enabling programmable post-transcriptional control. By mapping in pathogenic Escherichia coli the RNA interactome of CsrA, a global post-transcriptional regulator, we identify ~800 ternary complexes in which CsrA simultaneously binds an sRNA and an mRNA. Focusing on one class of these complexes, we show that PasE, a newly discovered sRNA, directs CsrA to virulence-associated mRNAs that are otherwise not targeted by CsrA, resulting in their repression. Altering the PasE seed sequence redirects CsrA to selected mRNAs, establishing a modular RNA-guided platform for gene regulation. Together, our findings reveal a new and likely widespread mechanism of bacterial post-transcriptional regulation and provide a framework for a new type of RNA-based synthetic regulation.

microbiology↗

bioRxiv2026

Bacterial nanotubes extend from a tubeosome organelle to establish an intercellular bridge

Bacterial nanotubes are widespread intercellular membranous bridges mediating molecular exchange among neighboring cells, yet their formation and dynamics remain largely unexplored. By combining live-cell imaging with in situ cryo-electron tomography, we describe nanotube biogenesis in the Gram-positive Bacillus subtilis. Nanotubes form elaborate, highly dynamic networks, with extending nanotubes exhibiting multidirectional movement toward nearby cells, repeatedly scanning their surface. Ultimately, nanotubes penetrate through the recipient cell wall to establish an intercellular bridge. Strikingly, nanotubes emerge within milliseconds from lipid-enriched sites representing a previously unrecognized membranous organelle, termed tubeosome, located in an unusual pseudo-periplasmic space beneath the cell wall. We present evidence that tubeosomes comprise proto-nanotube reservoirs that unfold to project nanotubes. Our study uncovers the tubeosome as a novel bacterial organelle and provides the first real-time visualization of intercellular bridge formation. As connecting nanotubes are found across bacteria, archaea, and eukaryotes, this mechanism may reflect a broadly conserved strategy for cellular communication.

microbiology↗

bioRxiv2025

A family of endonucleases that block nanotube-mediated plasmid dissemination

Small non-conjugative plasmids constitute a substantial portion of the bacterial mobile genome, driving the dissemination of beneficial genes, with their transfer primarily attributed to transformation, transduction, or co-mobilization with conjugative elements1-3. Here we explore an understudied plasmid spread route among bacteria, mediated by intercellular membranous nanotube conduits4. We reveal that, unlike traditional donor-to-recipient delivery, Nanotube-dependent Plasmid exchange (NPex) operates bidirectionally, enabling both plasmid donation and, to a lesser extent, plasmid acquisition. By identifying a Bacillus subtilis natural isolate deficient in NPex, we discovered a prophage-encoded factor, YokF, that blocks plasmid transmission, chiefly acting within the donor cell to inhibit plasmid donation. YokF is a nuclease that localizes to the membrane, where it interacts with a nanotube component to selectively impede plasmid transfer through degradation. Importantly, YokF homologs from various bacterial species were found to exhibit anti-NPex activity. Given their prevalence in Gram-positive bacteria, we propose that YokF homologs represent a conserved family of NPex gatekeepers that restrict plasmid flow within bacterial communities.

microbiology↗

bioRxiv2024

A conjugative plasmid exploits flagella rotation as a cue to facilitate its transfer

Conjugation-mediated DNA delivery is the primary mode for antibiotic resistance spread; yet, molecular mechanisms regulating the process remain largely unexplored. While conjugative plasmids typically rely on solid surfaces to facilitate donor-to-recipient proximity, the pLS20 conjugative plasmid, prevalent among Gram-positive Bacillus spp., uniquely requires fluid environments to motivate its transfer. Here we unveiled that pLS20, carried by B. subtilis, induces adhesin-promoted multicellular clustering, which can accommodate various species, offering a stable platform for DNA delivery in liquid milieu. We further discovered that induction of pLS20 promoters, governing crucial conjugative genes, hinges on the presence of donor cell flagella, the major bacterial motility organelle. Moreover, pLS20 regulatory circuit is strategically integrated into a mechanosensing signal transduction pathway responsive to flagella rotation, harnessing propelled flagella to activate conjugation genes exclusively during the host motile phase. This flagella-conjugation coupling strategy, provides the plasmid with the benefit of disseminating into remote destinations, infiltrating new niches.

microbiology↗

bioRxiv2024

EspH utilizes phosphoinositide and Rab binding domains to interact with plasma membrane infection sites and Rab GTPases

Enteropathogenic E. coli (EPEC) is a Gram-negative bacterial pathogen that causes persistent diarrhea. Upon attachment to the apical plasma membrane of the intestinal epithelium, the pathogen translocates virulent proteins called effectors into the infected cells. These effectors hijack numerous host processes for the pathogens benefit. Therefore, studying the mechanisms underlying their action is crucial for a better understanding of the disease. We show that translocated EspH interacts with multiple host Rab GTPases. AlphaFold predictions and site-directed mutagenesis identified glutamic acid and lysine at positions 37 and 41 as Rab interacting residues. Mutating these sites abolished the EspH ability to inhibit the Akt and mTORC1 signaling, lysosomal exocytosis, and bacterial invasion. Knocking out the endogenous Rab8a gene expression highlighted the involvement of Rab8a in Akt/mTORC1 signaling and lysosomal exocytosis. A phosphoinositide binding domain with a critical tyrosine was identified in EspH. Mutating the tyrosine abolished the localization of EspH at infection sites and its capacity to interact with Rabs. Our data suggest novel EspH-dependent mechanisms that elicit immune signaling and membrane trafficking during EPEC infection.

cell biology↗

bioRxiv2020

Activation of the type 3 secretion system of enteropathogenic E. coli leads to reprogrammingof its lipid metabolism

The cell envelope of gram-negative bacteria is a complex structure, essential for bacterial survival and for resistance to many antibiotics. Channels that cross the bacterial envelope and the host cell membrane form secretion systems that are activated upon attachment to host, enabling bacteria to inject effector molecules into the host cell, required for bacterial-host interaction. The type III secretion system (T3SS) is critical for the virulence of several pathogenic bacteria, including enteropathogenic E. coli (EPEC). The EPEC T3SS activation is associated with repression of carbon storage regulator (CsrA), resulting in gene expression remodeling, which is known to affect EPEC central carbon metabolism and contributes to the adaptation to a cell-adherent lifestyle in a poorly understood manner. We reasoned that the changes in bacterial envelope upon attachment to host and the activation of a secretion system may involve a modification of the lipid composition of bacterial envelope. Accordingly, we performed a lipidomics analysis on mutant strains that simulate T3SS activation. We saw a shift in glycerophospholipid metabolism towards the formation of lysophospholipids, attributed to corresponding upregulation of the phospholipase pldA and the acyltransferase ygiH upon T3SS activation in EPEC. We also detected a shift from menaquinones and ubiquinones to undecaprenyl lipids, concomitant to abnormal synthesis of O-antigen. The remodeling of lipid metabolism is mediated by CsrA and associated with increased bacteria cell size and Zeta potential, and a corresponding alteration in EPEC permeability to vancomycin, increasing the sensitivity of T3SS-activated strains and of adherent wild type EPEC to the antibiotic. ImportanceThe characterization of EPEC membrane lipid metabolism upon attachment to host is an important step towards a better understanding the shift of EPEC, a notable human pathogen, from a planktonic to adherent life style. It may also apply to other pathogenic bacteria that use this secretion system. We predict that upon attachment to host cells the lipid remodeling upon T3SS activation contributes to bacterial fitness and promotes host colonization, and show that it is associated with increased cell permeability and higher sensitivity to vancomycin. To the best of our knowledge, this is the first demonstration of a bacterial lipid remodeling due to activation of a secretion system.

microbiology↗