Biology subjects

Halliday, N.

Publications and source records attributed to Halliday, N..

5 recordsLinked to original sources

bioRxiv2024

The Quorum Sensing regulated sRNA Lrs1 is involved in the adaptation to low iron in Pseudomonas aeruginosa

Iron is an essential nutrient for microbial growth. The opportunistic pathogen Pseudomonas aeruginosa can survive under diverse conditions, including iron-depleted environments with the aid of small non-coding RNAs (sRNAs). P. aeruginosa also uses three quorum sensing (QS) systems Las, Rhl, and Pqs to coordinate virulence and infection establishment at the population level. This study links the sRNA Lrs1, which is located within the promoter of the Pqs biosynthetic operon pqsABCDE, to iron uptake regulation in the P. aeruginosa strain PAO1-L. Transcriptomics and phenotypic assays indicate that Lrs1 downregulates the production of the siderophore pyochelin but not pyoverdine, and that lrs1 regulation itself is dependent on iron availability. Although Lrs1 has been implicated in a positive feedback loop with the transcriptional regulator LasR in the strain PA14, the present findings indicate that this is not the case in PAO1-L in the tested conditions. Transcription of Lrs1 is dependent on quorum sensing, predominantly on RhlR with an auxiliary effect by PqsE. Furthermore, the Pqs system and phenazine production are modulated by Lrs1 only under iron limitation. This study identifies Lrs1 as a new QS-dependent post-transcriptional regulator of iron uptake and virulence highlighting its importance in environmental adaptation in P. aeruginosa. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=74 SRC="FIGDIR/small/627364v1_ufig1.gif" ALT="Figure 1"> View larger version (12K): org.highwire.dtl.DTLVardef@1585ca5org.highwire.dtl.DTLVardef@2a7d55org.highwire.dtl.DTLVardef@18a712eorg.highwire.dtl.DTLVardef@c84d2_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

bioRxiv2022

CTLA-4 regulates PD-L1-PD-1 interactions via transendocytosis of CD80

CTLA-4 and PD-1 are key immune checkpoints that are targeted in the treatment of cancer. Recently it has emerged that there is a physical interaction between the ligands of these pathways (CD80 and PD-L1), which can prevent PD-L1 inhibitory functions. Since CTLA-4 captures and degrades its ligands via transendocytosis, we investigated how transendocytosis of CD80 is impacted by PD-L1 interaction. We find that transendocytosis of CD80 results in a time-dependent recovery of PD-L1 availability that correlates with CD80 removal. Moreover, CD80 transendocytosis is highly specific in that only CD80 is removed, and its heterodimeric PD-L1 partner remains on the APC. We found no evidence that CTLA-4 interactions with CD80 were inhibited by PD-L1, however efficient removal of CD80 required an intact CTLA-4 cytoplasmic domain, distinguishing this process from more general trogocytosis. We also show that simple binding of CTLA-4 to the CD80-PD-L1 heterodimer is insufficient to liberate PD-L1-PD-1 interactions, suggesting that transendocytosis of CD80 is required to effectively control PD-L1-PD-1 interactions.

immunology↗

bioRxiv2022

Culture and genomic analysis of Symbiopectobacterium purcellii, gen.nov. sp. nov., isolated from the leafhopper Empoasca decipiens

Bacterial endosymbionts are found in multiple arthropod species, where they play crucial roles as nutritional symbionts, defensive symbionts or reproductive parasites. Recent work has highlighted a new clade of heritable microbes within the gammaproteobacteria that enter into both obligate and facultative symbioses, with an obligately required unculturable symbiont recently given the name Cand. Symbiopectobacterium. In this study, we describe a culturable rod shaped non-flagellated bacterial symbiont from this clade isolated from the leafhopper Empoasca decipiens. The symbiont is related to the transovarially-transmitted BEV bacterium that was first isolated from the leafhopper Euscelidius variegatus by Alexander Purcell, and we therefore name the symbiont Symbiopectobacterium purcellii sp. nov. gen. nov. We further report the closed genome sequence for S. purcellii. The genome is atypical for a heritable microbe, being large in size, without profound AT bias and with little evidence of pseudogenization. The genome is predicted to encode Type II, III and VI secretion systems and associated effectors and a non-ribosomal peptide synthase array likely to produce bioactive small molecules. Predicted metabolism is more complete than for other symbionts in the Symbiopectobacterium clade, and the microbe is predicted to synthesize a range of B vitamins. However, Biolog plate analysis indicate metabolism is depauperate compared to the sister clade, represented by Pectobacterium carotovorum. A quorum-sensing pathway related to that of Pectobacterium spp. (containing an overlapping expI-expR1 pair in opposite directions and a "solo" expR2) is evidenced, and LC-MS/MS analysis reveals the presence of 3-hydroxy-C10-HSL as the sole N-acylhomoserine lactone (AHL) in our strain. This AHL profile is profoundly divergent from that of other Erwinia and Pectobacterium spp., which produce mostly 3-oxo-C6- and 3-oxo-C8-HSL and could aid group identification. Thus, this microbe denotes one that has lost certain pathways associated with a saprophytic lifestyle but represents an important baseline against which to compare other members of the genus Symbiopectobacterium that show more profound integration into host biology.

microbiology↗

bioRxiv2021

Disruption of the Pseudomonas aeruginosa Tat system perturbs PQS-dependent quorum sensing and biofilm maturation through loss of the Rieske cytochrome bc1 sub-unit

Extracellular DNA (eDNA) is a major constituent of the extracellular matrix of Pseudomonas aeruginosa biofilms and its release is regulated via the pseudomonas quinolone signal (PQS) dependent quorum sensing (QS). By screening a P. aeruginosa transposon library to identify factors required for DNA release, mutants with insertions in the twin-arginine translocation (Tat) pathway were identified as exhibiting reduced eDNA release, and defective biofilm architecture with enhanced susceptibility to tobramycin. P. aeruginosa tat mutants showed substantial reductions in pyocyanin, rhamnolipid and membrane vesicle (MV) production consistent with perturbation of 2-heptyl-3-hydroxy-4-quinolone (PQS) dependent QS as demonstrated by changes in pqsA expression and 2-alkyl-4-quinolone (AQ) production. Provision of exogenous PQS to the tat mutants did not return pqsA, rhlA or phzA1 expression or pyocyanin production to wild type levels. However, transformation of the tat mutants with the AQ-independent pqs effector pqsE restored phzA1 expression and pyocyanin production. Since mutation or inhibition of Tat prevented PQS-driven auto-induction, we sought to identify the Tat secretion substrate responsible. A pqsA::lux fusion was introduced into each of 34 validated P. aeruginosa Tat substrate deletion mutants. Analysis of each mutant for reduced bioluminescence revealed that the signalling defect was associated with the Rieske iron-sulfur subunit of the cytochrome bc1 complex. In common with the parent strain, a Rieske mutant exhibited defective PQS signalling, AQ production, rhlA expression and eDNA release that could be restored by genetic complementation. Thus, lack of the Rieske sub-unit export is clearly responsible for the Tat-mediated perturbation of PQS-dependent QS, the loss of virulence factor production, biofilm eDNA and the tobramycin tolerance of P. aeruginosa biofilms. Author SummaryPseudomonas aeruginosa is a highly adaptable human pathogen responsible for causing chronic biofilm-associated infections. Biofilms are highly refractory to host defences and antibiotics and thus difficult to eradicate. The biofilm extracellular matrix incorporates extracellular DNA (eDNA). This stabilizes biofilm architecture and helps confer tolerance to antibiotics. Since mechanisms that control eDNA release are not well understood, we screened a P. aeruginosa mutant bank for strains with defects in eDNA release and discovered a role for the twin-arginine translocation (Tat) pathway that exports folded proteins across the cytoplasmic membrane. Perturbation of the Tat pathway resulted in defective biofilms susceptible to antibiotic treatment as a consequence of perturbed pseudomonas quinolone (PQS) signalling. This resulted in the failure to produce or release biofilm components including eDNA, phenazines and rhamnolipids as well as microvesicles. Furthermore, we discovered that perturbation of PQS signalling was a consequence of the inability of tat mutants to translocate the Rieske subunit of the cytochrome bc1 complex involved in electron transfer and energy transduction. Given the importance of PQS signalling and the Tat system to virulence and biofilm maturation in P. aeruginosa, our findings underline the potential of the Tat system as a drug target for novel antimicrobial agents.

microbiology↗

bioRxiv2019

Cross-kingdom signalling regulates spore germination in the moss Physcomitrella patens

Plants live in close association with microorganisms that can have beneficial or detrimental effects. The activity of bacteria in association with flowering plants has been extensively analysed. Bacteria use quorum-sensing as a way of monitoring their population density and interacting with their environment. A key group of quorum sensing molecules in Gram-negative bacteria are the N-acylhomoserine lactones (AHLs), which are known to affect the growth and development of both flowering plants, including crops, and marine algae. Thus, AHLs have potentially important roles in agriculture and aquaculture. Nothing is known about the effects of AHLs on the earliest-diverging land plants, thus the evolution of AHL-mediated bacterial-plant- and algal interactions is unknown. In this paper, we show that AHLs can affect spore germination in a representative of the earliest plants on land, the Bryophyte moss Physcomitrella patens. Furthermore, we demonstrate that sporophytes of wild isolates of Physcomitrella patens are associated with AHL-producing bacteria.

plant biology↗