Biology subjects

Pastora, A. B.

Publications and source records attributed to Pastora, A. B..

3 recordsLinked to original sources

bioRxiv2024

Multiple Pathways Impact Swarming Motility of Pseudomonas fluorescens Pf0-1

Swarming motility in pseudomonads typically requires both a functional flagellum and production/secretion of a biosurfactant. Published work has shown that the wild-type Pseudomonas fluorescens Pf0-1 is swarming-deficient due to a point mutation in the gacA gene, which until recently, was thought to inactivate rather than attenuate the Gac/Rsm pathway. As a result, little is known about the underlying mechanisms that regulate swarming motility by P. fluorescens Pf0-1. Here, we demonstrate that a {Delta}rsmA {Delta}rsmE {Delta}rsmI mutant, which phenotypically mimics Gac/Rsm pathway overstimulation, is proficient at swarming motility. RsmA and RsmE appear to play a key role in this regulation. Transposon mutagenesis of the {Delta}rsmA {Delta}rsmE {Delta}rsmI mutant identified multiple factors that impact swarming motility, including pathways involved in flagellar synthesis and biosurfactant production/secretion. We find that loss of genes linked to biosurfactant Gacamide A biosynthesis or secretion impact swarming motility, as does loss of the alternative sigma factor FliA, which results in a defect in flagellar function. Collectively, these findings provide evidence that P. fluorescens Pf0-1 can swarm if the Gac/Rsm pathway is activated, highlight the regulatory complexity of swarming motility in this strain, and demonstrate that the cyclic lipopeptide Gacamide A is utilized as a biosurfactant for swarming motility. ImportanceSwarming motility is a coordinated process that allows communities of bacteria to collectively move across a surface. For P. fluorescens Pf0-1, this phenotype is notably absent in the parental strain and to date little is known about the regulation of swarming in this strain. Here, we identify RsmA and RsmE as key repressors of swarming motility via modulating the levels of biosurfactant production/secretion. Via transposon mutagenesis and subsequent genetic analyses, we further identify potential regulatory mechanisms of swarming motility and link Gacamide A biosynthesis and transport machinery to swarming motility.

microbiology↗

bioRxiv2023

The Regulator FleQ Post-Transcriptionally Regulates the Production of RTX Adhesins by Pseudomonas fluorescens

Biofilm formation by the Gram-negative gammaproteobacterium Pseudomonas fluorescens relies on the production of the repeat-in-toxin (RTX) adhesins LapA and MapA in the cytoplasm, secretion of these adhesins through their respective type 1 secretion systems, and retention at the cell surface. Published work has shown that retention of the adhesins occurs via a post-translational mechanism involving the cyclic-di-GMP receptor LapD and the protease LapG. However, little is known about the underlying mechanisms that regulate the production of these adhesins. Here, we demonstrate that the master regulator FleQ modulates biofilm formation by post-transcriptionally regulating the production of LapA and MapA. We find that a {Delta}fleQ mutant has a biofilm formation defect compared to the WT strain, which is attributed in part to a decrease in LapA and MapA production, despite the {Delta}fleQ mutant having increased levels of lapA and mapA transcripts compared to the WT strain. Through transposon mutagenesis and subsequent genetic analysis, we found that over-stimulation of the Gac/Rsm pathway partially rescues biofilm formation in the {Delta}fleQ mutant background. Collectively, these findings provide evidence that FleQ regulates biofilm formation by post-transcriptionally regulating the production of LapA and MapA, and that activation of the Gac/Rsm pathway can enhance biofilm formation by P. fluorescens. ImportanceBiofilm formation is a highly coordinated process that bacteria undergo to colonize a variety of surfaces. For Pseudomonas fluorescens, biofilm formation requires the production and localization of RTX adhesins to the cell surface. To date, little is known about the underlying mechanisms that regulate biofilm formation by P. fluorescens. Here, we identify FleQ as a key regulator of biofilm formation that modulates the production of LapA and MapA through a post-transcriptional mechanism. We provide further evidence implicating activation of the Gac/Rsm system in FleQ-dependent regulation of biofilm formation. Together, our findings uncover evidence for a mechanism of post-transcriptional regulation of the LapA/MapA adhesins.

microbiology↗

bioRxiv2020

MapA, a second large RTX adhesin, contributes to biofilm formation by Pseudomonas fluorescens

Mechanisms by which cells attach to a surface and form a biofilm are diverse and differ greatly between organisms. The Gram-negative, Gammaproteobacterium Pseudomonas fluorescens attaches to a surface through the localization of the large type 1-secreted RTX adhesin LapA to the outer surface of the cell. LapA localization to the cell surface is controlled by the activities of a periplasmic protease, LapG and an inner-membrane spanning cyclic di-GMP responsive effector protein, LapD. A previous study identified a second, LapA-like protein encoded in the P. fluorescens Pf0-1 genome: Pfl01_1463. However, deletion of this gene had no discernible phenotype under our standard laboratory growth conditions. Here, we identified specific growth conditions wherein, Pfl01_1463, hereafter called MapA (Medium Adhesion Protein A) is a functional adhesin contributing to biofilm formation. This adhesin, like LapA, appears to be secreted through a Lap-related type 1 secretion machinery. We show MapA involvement in biofilm formation is also controlled by LapD and LapG, and that the differing roles of LapA and MapA in biofilm formation are achieved, at least in part, through the differences in the sequences of the two adhesins and their differential, cyclic di-GMP-dependent transcriptional regulation. This differential regulation appears to lead to different distributions of the expression of lapA and mapA within a biofilm. Our data indicate that the mechanisms by which a cell forms a biofilm and the components of a biofilm matrix can differ depending on growth conditions in the biofilm. ImportanceAdhesins are critical for the formation and maturation of bacterial biofilms. We identify a second adhesin in P. fluorescens, called MapA, which appears to play a role in biofilm maturation and whose regulation is distinct from the previously reported LapA adhesin, which is critical for biofilm initiation. Analysis of bacterial adhesins show that LapA-like and MapA-like adhesins are found broadly in Pseudomonads and related organisms, indicating that the utilization of different suites of adhesins may be broadly important in the Gammaproteobacteria.

microbiology↗