Project description:Biofilms are surface-adhered bacterial communities encased in an extracellular matrix composed of polysaccharides, proteins, and extracelluar (e)DNA, with eDNA being required for the formation and integrity of biofilms. Here we demonstrate that the spatial and temporal release of eDNA is regulated by BfmR, a regulator essential for Pseudomonas aeruginosa biofilm development. The expression of bfmR coincided with localized cell death and DNA release, with high eDNA concentrations localized to the outer part of microcolonies in the form of a ring and as a cap on small clusters. Additionally, eDNA release and cell lysis increased significantly following bfmR inactivation. Genome-wide transcriptional profiling indicated that bfmR was required for repression of genes associated with bacteriophage assembly and bacteriophage-mediated lysis. In order to determine which of these genes were directly regulated by BfmR, we utilized chromatin immunoprecipitation (ChIP) analysis to identify the promoter of PA0691, termed here phdA, encoding a previously undescribed homologue of the prevent-host-death (Phd) family of proteins. Lack of phdA expression coincided with impaired biofilm development, increased cell death and bacteriophage release, a phenotype comparable to ΔbfmR. Expression of phdA in ΔbfmR biofilms restored eDNA release, cell lysis, release of bacteriophages, and biofilm formation to wild type levels. Moreover, overexpression of phdA rendered P. aeruginosa resistant to lysis mediated by superinfective bacteriophage Pf4 which was only detected in biofilms. The expression of bfmR was stimulated by conditions resulting in membrane perturbation and cell lysis. Thus, we propose that BfmR regulates biofilm development by controlling bacteriophage-mediated lysis and thus, cell death and eDNA release, via PhdA.
Project description:We examined gene expression profiling of native mussels that were sampled in early summer 2003 from sites of the Venice lagoon area known to be differently affected by chemical pollution: Sites 1 and 2 close to the industrial district of Marghera and Site 3 close to the Lido lagoon outlet. Site 4, a current mussel farm located offshore, has been chosen as source of reference targets for microarray hybridizations. We have limited the preliminary assessment to the digestive gland. Digestive gland total RNA of each Site was hybridized in competition with the offshore mussels (Site 4 - Reference) and the relative abundance of each gene was measured by directly comparing fluorescent signals for each probe. We carried out two separate hybridizations for each site of the Venice lagoon area.. Keywords = digestive gland Keywords = Venice lagoon Keywords = chemical pollution Keywords = native mussels Keywords = transcriptional profiling Keywords: ordered
Project description:Ribosome biogenesis is essential for protein synthesis in gene expression. Yeast eIF5B has been shown biochemically to facilitate 18S rRNA 3’ end maturation during late-40S ribosomal subunit assembly and gate the transition from translation initiation to elongation. But the effects of eIF5B have not been studied at the genome-wide level in any organism, and 18S rRNA 3’ end maturation is poorly understood in plants. Arabidopsis HOT3/eIF5B1 was found to promote development and heat-stress acclimation by translational regulation, but its molecular function remained unknown. Here, we show that HOT3 is a late-stage ribosome biogenesis factor that facilitates 18S rRNA 3’ end processing and is a translation initiation factor that globally impacts the transition from initiation to elongation. By developing and implementing 18S-ENDseq, we revealed previously unknown events in 18S rRNA 3’ end maturation or metabolism. We quantitatively defined new processing hotspots and identified adenylation as the prevalent non-templated RNA modification at the 3’ ends of pre-18S rRNAs. Aberrant 18S rRNA maturation in hot3 further activated RNAi to generate RDR1- and DCL2/4-dependent risiRNAs mainly from a 3’ portion of 18S rRNA. We further showed that risiRNAs in hot3 were predominantly localized in ribosome-free fractions not responsible for the 18S rRNA maturation or translation initiation defects in hot3. Our study uncovered the molecular function of HOT3/eIF5B1 in 18S rRNA maturation at the late-40S assembly stage and revealed the regulatory crosstalk among ribosome biogenesis, mRNA translation initiation, and siRNA biogenesis in plants.
Project description:We identify a group of bacterial genes that are induced and repressed by the addition of eDNA, due to cation chelation, acidification or nutrient utilization.
Project description:Monitoring microbial communities can aid in understanding the state of these habitats. Environmental DNA (eDNA) techniques provide efficient and comprehensive monitoring by capturing broader diversity. Besides structural profiling, eDNA methods allow the study of functional profiles, encompassing the genes within the microbial community. In this study, three methodologies were compared for functional profiling of microbial communities in estuarine and coastal sites in the Bay of Biscay. The methodologies included inference from 16S metabarcoding data using Tax4Fun, GeoChip microarrays, and shotgun metagenomics.