Project description:The stringent stress response, mediated by the signalling molecule guanosine penta(tetra)-phosphate (p)ppGpp, has recently gained more attraction as a potential new therapeutic target. However, interference of a global mediator could lead to unwanted side effects, therefore warrants detailed mechanistic understanding. We used RNA-Seq to investigate the transcriptional landscape of a Pseudomonas aeruginosa ppGpp-deficient strain under non-stressful conditions. In exponential growth phase in nutrient-rich broth, the lack of ppGpp led to >1600 transcriptional changes, which provided a novel aspect of the importance of ppGpp in general rather than having the traditional stress/starvation induced changes. Intracellular protein tracking indicated that RelA was bound to the ribosome while SpoT localized at the cell poles. Commonly dysregulated functional systems were divided into 34 clusters and revealed individual genes that might belong to the same pathway as other known adjacent regulon members. The regulatory network of ppGpp included global transcriptional regulators AlgR, OxyR, LasR, and AmrZ that further indirectly linked the stress response to secretion systems and polysaccharide production. The stress response directly influenced the expression of quorum sensing and other downstream effectors including AMB toxin production, the ECF-sigma factor PvdS, the transcriptional regulator PtxS, the two-component response regulator PprB, and various proteases such as ImpA, PA0572, ClpP2, and AprA.
Project description:Transcriptomic and proteomic insights into innate immunity and adaptations to a symbiotic lifestyle in the gutless marine worm Olavius algarvensis
Project description:Unicellular algae, termed phytoplankton, greatly impact the marine environment by serving as the basis of marine food webs and by playing central roles in biogeochemical cycling of elements. The interactions between phytoplankton and heterotrophic bacteria affect the fitness of both partners. It is becoming increasingly understood that metabolic exchange determines the nature of such interactions, but the underlying molecular mechanisms remain underexplored. Here, we investigated the molecular and metabolic basis for the bacterial lifestyle switch, from coexistence to pathogenicity, in Sulfitobacter D7 during interactions with Emiliania huxleyi, a cosmopolitan bloom-forming phytoplankter. The interaction displays two distinct phases: first, there is a coexisting phase in which the alga grows exponentially and the bacterium grows as well. The interaction shifts to pathogenic when the virulence of Sulfitobacter D7 towards E. huxleyi is invoked upon exposure to high concentrations of algal dimethylsulfoniopropionate (DMSP), which occurs when the algae reach stationary growth or when DMSP is applied exogenously to algae in exponential growth. We aimed to unravel the response of Sulfitobacter D7 to the pathogenicity-inducing compound, DMSP, and to different algae-derived infochemicals that affect the lifestyle of the bacterium. We grew Sulfitobacter D7 in conditioned media (CM) derived from algal cultures at the different growth phases, exponential and stationary (Exp-CM and Stat-CM, respectively), in which DMSP concentration is low and high, respectively. This enabled us to separate between different phases of the interaction with E. huxleyi, i.e., Exp-CM representing the coexisting phase, and Stat-CM representing the pathogenic phase. An additional pathogenicity-inducing treatment was Exp-CM supplemented with 100 µM DMSP (herein Exp-CM+DMSP). This condition mimicked co-cultures to which we added DMSP exogenously and thus induced Sulfitobacter D7 pathogenicity, which lead to death of exponentially growing E. huxleyi. In order to identify bacterial genes that are specifically responsive to DMSP, and are not affected by other algae-derived factors, we grew Sulfitobacter D7 in defined minimal medium (MM), lacking algal metabolites, supplemented with 100 µM DMSP (herein MM+DMSP), and examined the transcriptional response. After 24 h of Sulfitobacter D7 growth in all 5 media, triplicates were taken for transcriptomic analysis. Altogether, this experimental design allowed to expand our understanding on the bacterial response to DMSP, algal infochemicals and which of these are essential for coexistence and pathogenicity.
Project description:Salmonella enterica serovar Typhimurium (S. Typhimurium) is a zoonotic pathogen that causes diarrheal disease in humans and animals. During salmonellosis, S. Typhimurium colonizes epithelial cells lining the gastrointestinal tract. S. Typhimurium has an unusual lifestyle in epithelial cells that begins within an endocytic-derived Salmonella-containing vacuole (SCV), followed by escape into the cytosol, epithelial cell lysis and bacterial release. The cytosol is a more permissive environment than the SCV and supports rapid bacterial growth. The physicochemical conditions encountered by S. Typhimurium within the cytosol, and the bacterial genes required for cytosolic colonization, remain unknown. Here we have exploited the parallel colonization strategies of S. Typhimurium in epithelial cells to decipher the two niche-specific bacterial virulence programs. By combining a population-based RNA-seq approach with single-cell microscopic analysis, we identified bacterial genes/sRNAs with cytosol-specific or vacuole-specific expression signatures. Using these genes/sRNAs as environmental biosensors, we defined that Salmonella is exposed to iron and manganese deprivation and oxidative stress in the cytosol and zinc and magnesium deprivation in the SCV. Furthermore, iron availability was critical for optimal S. Typhimurium replication in the cytosol, as well as entC, fepB, soxS and sitA-mntH. Virulence genes that are typically associated with extracellular bacteria, namely Salmonella pathogenicity island 1 (SPI1) and SPI4, had a cytosolic-specific expression profile. Our study reveals that the cytosolic and vacuolar S. Typhimurium virulence gene programs are unique to, and tailored for, residence within distinct intracellular compartments. Therefore, this archetypical vacuole-adapted pathogen requires extensive transcriptional reprogramming to successfully colonize the mammalian cytosol.
Project description:Epigenome analysis of 18 subjects and 20 controls, before, midpoint, and after a change in diet, exercise, and other lifestyle modifications
Project description:Analysis of breast cancer survivors' gut microbiota after lifestyle intervention, during the COVID-19 lockdown, by 16S sequencing of fecal samples.
Project description:To investigate the microRNAs involved in the processes of beneficial and detrimental lifestyles, including caloric restriction(CR), exercise and high-fat diet(HF), we performed a comprehensive and thorough comparison of microRNAs expression profiles in liver among these lifestyle modifications.
Project description:The objective of this study was to examine relationships between weight loss through changes in lifestyle and peripheral blood gene expression profiles. Substantial weight loss (-15.2+3.8%) in lifestyle participants was associated with improvement in selected cardiovascular risk factors and significant changes in peripheral blood gene expression from pre- to post-intervention: 132 unique genes showed significant expression changes related to immune function and inflammatory responses involving endothelial activation. In contrast, participants losing minimal weight (-3.1+2.5%) showed only minor changes in cardiovascular risk factors and markers of inflammation, and no changes in gene expression compared to non-intervention controls after 1 year. Weight loss (>10%) during lifestyle modification is associated with down-regulation of genetic pathways governing interactions between circulating immune cells and the vascular endothelium and may be required to successfully reduce CVD risk. A prospective nonrandomized trail was conducted over 1 year in participants undergoing intensive lifestyle modification to reverse or stabilize progression of coronary artery disease. Cardiovascular risk factors, inflammatory biomarkers, and gene expression as a function of weight loss were assessed in 89 lifestyle participants and 71 retrospectively matched controls undergoing usual care.