Biology subjects

Monlezun, L.

Publications and source records attributed to Monlezun, L..

4 recordsLinked to original sources

bioRxiv2026

Tvp complexes support formation of the VgrG-PAAR spike during Type VI secretion system assembly

Type VI secretion systems (T6SSs) are nanomachineries used by bacteria to inject toxic effectors into neighbouring cells during interbacterial competition or infection. The existence of multiple paralogues of the structural proteins VgrG and PAAR and of specific accessory proteins allows the same T6SS to deliver a wide range of effectors. We describe a new set of accessory proteins required for assembly of T6SSs containing the VgrG1-Paar1 spike and delivery of a novel VgrG1-associated membrane-targeting effector in Serratia marcescens. TvpAB, TvpB, TvpC and Paar1 form a pre-complex essential for T6SS assembly, with VgrG1 subsequently replacing TvpC. Phylogenetic analysis and structural modelling reveal that Tvp proteins are widely conserved but Tvp-containing complexes vary in organisation and complexity. We define three classes of Tvp complex, all sharing a DUF2169-containing TvpA protein which stabilises a DUF4150/PAAR-containing protein. Class I, which includes only TvpA, functions without a pre-complex, whereas Classes II and III involve additional Tvp components and two-step assembly. Our findings highlight how modular effector recruitment strategies underlie the versatility of the T6SS and suggest how alternative Tvp systems are tailored to promote assembly of secretion-competent, effector-loaded VgrG-PAAR spikes.

microbiology↗

bioRxiv2024

Base composition at the start of the coding sequence controls the balance between translation initiation and mRNA degradation in E. coli .

Protein synthesis efficiency is highly dependent on the mRNA coding sequence. Furthermore, there is extensive evidence of a correlation between mRNA stability and protein expression level, though the mechanistic determinants remain unclear. Using yellow fluorescent protein (YFP) as a reporter gene, we herein demonstrate that adenosine (A) abundance in the first six codons is a critical determinant for achieving high protein synthesis in E. coli. Increasing A and/or decreasing guanosine (G) content in this region with synonymous codons results in substantial increases in protein expression level both in vivo and in vitro that are correlated with steady-state mRNA concentration in vivo. The change in mRNA concentration is attributable to changes in the stability of the mRNA that are directly coupled to its translation efficiency. Increasing A content promotes mRNA incorporation into the functional 70S ribosomal initiation complex without altering its affinity for the 30S ribosomal subunit. These results support a model in which base composition in the first six codons modulates local mRNA folding energy and single-strandedness to control the balance between productive translation initiation versus degradation of mRNAs bound to the 30S ribosomal subunit. Based on these findings, we developed a short N-terminal coding sequence that optimizes translation initiation efficiency for protein production in E. coli.

microbiology↗

bioRxiv2023

Global regulation via modulation of ribosome pausing by EttA

Having multiple rounds of translation of the same mRNA creates dynamic complexities along with opportunities for regulation related to ribosome pausing and stalling at specific sequences. Yet, mechanisms controlling these critical processes and the principles guiding their evolution remain poorly understood. Through genetic, genomic, physiological, and biochemical approaches, we demonstrate that regulating ribosome pausing at specific amino acid sequences can produce [~]2-fold changes in protein expression levels which strongly influence cell growth and therefore evolutionary fitness. We demonstrate, both in vivo and in vitro, that the ABC-F protein EttA directly controls the translation of mRNAs coding for a subset of enzymes in the tricarboxylic acid (TCA) cycle and its glyoxylate shunt, which modulates growth in some chemical environments. EttA also modulates expression of specific proteins involved in metabolically related physiological and stress-response pathways. These regulatory activities are mediated by EttA rescuing ribosomes paused at specific patterns of negatively charged residues within the first 30 amino acids of nascent proteins. We thus establish a unique global regulatory paradigm based on sequence-specific modulation of translational pausing.

microbiology↗

bioRxiv2019

A new family of Type VI secretion system-delivered effector proteins displays ion-selective pore-forming activity

Type VI secretion systems (T6SSs) are nanomachines widely used by bacteria to compete with rivals. T6SSs deliver multiple toxic effector proteins directly into neighbouring cells and play key roles in shaping diverse polymicrobial communities. A number of families of T6SS-dependent anti-bacterial effectors have been characterised, however the mode of action of others remains unknown. Here we report that Ssp6, an anti-bacterial effector delivered by the Serratia marcescens T6SS, is an ion-selective pore-forming toxin. In vivo, Ssp6 inhibits growth by causing depolarisation of the inner membrane of intoxicated cells and also leads to increased outer membrane permeability, whilst reconstruction of Ssp6 activity in vitro demonstrated that it forms cation-selective pores. A survey of bacterial genomes revealed that Ssp6-like effectors are widespread in Enterobacteriaceae and often linked with T6SS genes. We conclude that Ssp6 represents a new family of T6SS-delivered anti-bacterial effectors, further diversifying the portfolio of weapons available for deployment during inter-bacterial conflict.

microbiology↗