Unraveling Fe(II)-oxidizing mechanisms in a facultative Fe(II)-oxidizer, Sideroxydans lithotrophicus ES-1 via culturing, transcriptomics, and RT-qPCR

Sideroxydans lithotrophicus ES-1 grows autotrophically either by Fe(II) oxidation or thiosulfate oxidation, in contrast to most other neutrophilic Fe(II)-oxidizing bacteria (FeOB) isolates. This provides a unique opportunity to explore the physiology of a facultative FeOB and constrain the genes specific to Fe(II) oxidation. We compared the growth of S. lithotrophicus ES-1 on Fe(II), thiosulfate, and both substrates together. While initial growth rates were similar, thiosulfate-grown cultures had higher yield with or without Fe(II) present, which may give ES-1 an advantage over obligate FeOB. To investigate the Fe(II) and S oxidation pathways, we conducted transcriptomics experiments, validated with RT-qPCR. We explored the long-term gene expression response at different growth phases (over days-week) and expression changes during a short-term switch from thiosulfate to Fe(II) (90 min). The dsr and sox sulfur oxidation genes were upregulated in thiosulfate cultures. The Fe(II) oxidase gene cyc2 was among the top expressed genes during both Fe(II) and thiosulfate oxidation, and addition of Fe(II) to thiosulfate-grown cells caused an increase in cyc2 expression. These results support the role of Cyc2 as the Fe(II) oxidase and suggest that ES-1 maintains readiness to oxidize Fe(II) even in the absence of Fe(II). We used gene expression profiles to further constrain the ES-1 Fe(II) oxidation pathway. Notably, among the most highly upregulated genes during Fe(II) oxidation were genes for alternative complex III, reverse electron transport and carbon fixation. This implies a direct connection between Fe(II) oxidation and carbon fixation, suggesting that CO 2 is an important electron sink for Fe(II) oxidation. Importance Neutrophilic FeOB are increasingly observed in various environments, but knowledge of their ecophysiology and Fe(II) oxidation mechanisms is still relatively limited. Sideroxydans are widely observed in aquifers, wetlands, and sediments, and genome analysis suggests metabolic flexibility contributes to their success. The type strain ES-1 is unusual amongst neutrophilic FeOB isolates as it can grow on either Fe(II) or a non-Fe(II) substrate, thiosulfate. Almost all our knowledge of neutrophilic Fe(II) oxidation pathways comes from genome analyses, with some work on metatranscriptomes. This study used culture-based experiments to test the genes specific to Fe(II) oxidation in a facultative FeOB and refine our model of the Fe(II) oxidation pathway. We gained insight into how facultative FeOB like ES-1 connect Fe, S, and C biogeochemical cycling in the environment, and suggest a multi-gene indicator would improve understanding of Fe(II) oxidation activity in environments with facultative FeOB.

Download Full-text

Unraveling Fe(II)-oxidizing mechanisms in a facultative Fe(II) oxidizer Sideroxydans lithotrophicus ES-1 via culturing, transcriptomics, and RT-qPCR

10.1101/2021.08.18.456919 ◽

2021 ◽

Author(s):

Nanqing Zhou ◽

Jessica L. Keffer ◽

Shawn W. Polson ◽

Clara S Chan

Keyword(s):

Gene Expression ◽

Carbon Fixation ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Sulfur Oxidation ◽

Complex Iii ◽

Initial Growth ◽

Gene Expression Response ◽

Oxidase Gene ◽

Thiosulfate Oxidation

Download Full-text

Gene expression response of the rat small intestine following oralSalmonellainfection

Physiological Genomics ◽

10.1152/physiolgenomics.00190.2006 ◽

2007 ◽

Vol 30 (2) ◽

pp. 123-133 ◽

Cited By ~ 26

Author(s):

Wendy Rodenburg ◽

Ingeborg M. J. Bovee-Oudenhoven ◽

Evelien Kramer ◽

Roelof van der Meer ◽

Jaap Keijer

Keyword(s):

Gene Expression ◽

Expression Profiles ◽

Inflammatory Marker ◽

Molecular Response ◽

Lipocalin 2 ◽

Ileal Mucosa ◽

Gene Expression Response ◽

Expression Response

Data on the molecular response of the intestine to the food-borne pathogen Salmonella are derived from in vitro studies, whereas in vivo data are lacking. We performed an oral S. enteritidis infection study in Wistar rats to obtain insight in the in vivo response in time. Expression profiles of ileal mucosa (IM) and Peyer's patches (PP) were generated using DNA microarrays at days 1, 3, and 6 postinfection. An overview of Salmonella-regulated processes was obtained and confirmed by quantitative real-time PCR on pooled and individual samples. Salmonella-induced gene expression responses in vivo are fewer and smaller than observed in vitro, and the response develops over a longer period of time. Few effects are seen at day 1 and mainly occur in IM, suggesting the mucosa as the primary site of invasion. Later, a bigger response is observed, especially in PP. Decreased expression of anti-microbial peptides genes (in IM at day 1) suggests inhibition of this process by Salmonella. Newly identified target processes are carbohydrate transport (increased expression in IM at day 1) and phase I and phase II detoxification (decreased expression at days 3 and 6). Increase of cytokine and chemokine expression occurs at later time points, both in PP and IM. Pancreatitis-associated protein, lipocalin 2, and calprotectin, potential inflammatory marker proteins, showed induced expression from day 3 onward. We conclude that the in vivo gene expression response of the ileum to Salmonella differs to a large extent from the response seen in vitro.

Download Full-text

Similarities and differences between smoking-related gene expression in nasal and bronchial epithelium

Physiological Genomics ◽

10.1152/physiolgenomics.00167.2009 ◽

2010 ◽

Vol 41 (1) ◽

pp. 1-8 ◽

Cited By ~ 80

Author(s):

Xiaoling Zhang ◽

Paola Sebastiani ◽

Gang Liu ◽

Frank Schembri ◽

Xiaohui Zhang ◽

...

Keyword(s):

Gene Expression ◽

Respiratory Tract ◽

Cigarette Smoke ◽

Expression Profiles ◽

Physiological Responses ◽

Bronchial Epithelium ◽

Nasal Epithelium ◽

Cigarette Smoke Exposure ◽

Related Gene ◽

Gene Expression Response

Previous studies have shown that physiological responses to cigarette smoke can be detected via bronchial airway epithelium gene expression profiling and that heterogeneity in this gene expression response to smoking is associated with lung cancer. In this study, we sought to determine the similarity of the effects of tobacco smoke throughout the respiratory tract by determining patterns of smoking-related gene expression in paired nasal and bronchial epithelial brushings collected from 14 healthy nonsmokers and 13 healthy current smokers. Using whole genome expression arrays, we identified 119 genes whose expression was affected by smoking similarly in both bronchial and nasal epithelium, including genes related to detoxification, oxidative stress, and wound healing. While the vast majority of smoking-related gene expression changes occur in both bronchial and nasal epithelium, we also identified 27 genes whose expression was affected by smoking more dramatically in bronchial epithelium than nasal epithelium. Both common and site-specific smoking-related gene expression profiles were validated using independent microarray datasets. Differential expression of select genes was also confirmed by RT-PCR. That smoking induces largely similar gene expression changes in both nasal and bronchial epithelium suggests that the consequences of cigarette smoke exposure can be measured in tissues throughout the respiratory tract. Our findings suggest that nasal epithelial gene expression may serve as a relatively noninvasive surrogate to measure physiological responses to cigarette smoke and/or other inhaled exposures in large-scale epidemiological studies.

Download Full-text

Validating the Cyc2 Neutrophilic Iron Oxidation Pathway Using Meta-omics of Zetaproteobacteria Iron Mats at Marine Hydrothermal Vents

mSystems ◽

10.1128/msystems.00553-19 ◽

2020 ◽

Vol 5 (1) ◽

Cited By ~ 10

Author(s):

Sean M. McAllister ◽

Shawn W. Polson ◽

David A. Butterfield ◽

Brian T. Glazer ◽

Jason B. Sylvan ◽

...

Keyword(s):

Iron Oxides ◽

Hydrothermal Vents ◽

Carbon Fixation ◽

Marker Gene ◽

Iron Oxidation ◽

Oxidase Gene ◽

Content Type ◽

Oxidation Pathway ◽

Fe Oxidation ◽

Iron Oxidizers

ABSTRACT Zetaproteobacteria create extensive iron (Fe) oxide mats at marine hydrothermal vents, making them an ideal model for microbial Fe oxidation at circumneutral pH. Comparison of neutrophilic Fe oxidizer isolate genomes has revealed a hypothetical Fe oxidation pathway, featuring a homolog of the Fe oxidase Cyc2 from Acidithiobacillus ferrooxidans. However, Cyc2 function is not well verified in neutrophilic Fe oxidizers, particularly in Fe-oxidizing environments. Toward this, we analyzed genomes and metatranscriptomes of Zetaproteobacteria, using 53 new high-quality metagenome-assembled genomes reconstructed from Fe mats at Mid-Atlantic Ridge, Mariana Backarc, and Loihi Seamount (Hawaii) hydrothermal vents. Phylogenetic analysis demonstrated conservation of Cyc2 sequences among most neutrophilic Fe oxidizers, suggesting a common function. We confirmed the widespread distribution of cyc2 and other model Fe oxidation pathway genes across all represented Zetaproteobacteria lineages. High expression of these genes was observed in diverse Zetaproteobacteria under multiple environmental conditions and in incubations. The putative Fe oxidase gene cyc2 was highly expressed in situ, often as the top expressed gene. The cyc2 gene showed increased expression in Fe(II)-amended incubations, with corresponding increases in carbon fixation and central metabolism gene expression. These results substantiate the Cyc2-based Fe oxidation pathway in neutrophiles and demonstrate its significance in marine Fe-mineralizing environments. IMPORTANCE Iron oxides are important components of our soil, water supplies, and ecosystems, as they sequester nutrients, carbon, and metals. Microorganisms can form iron oxides, but it is unclear whether this is a significant mechanism in the environment. Unlike other major microbial energy metabolisms, there is no marker gene for iron oxidation, hindering our ability to track these microbes. Here, we investigate a promising possible iron oxidation gene, cyc2, in iron-rich hydrothermal vents, where iron-oxidizing microbes dominate. We pieced together diverse Zetaproteobacteria genomes, compared these genomes, and analyzed expression of cyc2 and other hypothetical iron oxidation genes. We show that cyc2 is widespread among iron oxidizers and is highly expressed and potentially regulated, making it a good marker for the capacity for iron oxidation and potentially a marker for activity. These findings will help us understand and potentially quantify the impacts of neutrophilic iron oxidizers in a wide variety of marine and terrestrial environments.

Download Full-text

179. PURIFICATION AND CHARACTERISATION OF MOUSE TESTICULAR MACROPHAGES: GENE EXPRESSION RESPONSE TO LIPOPOLYSACCHARIDE ACTIVATION INDICATES AN IMMUNOSUPPRESSIVE PHENOTYPE

Reproduction Fertility and Development ◽

10.1071/srb10abs179 ◽

2010 ◽

Vol 22 (9) ◽

pp. 97

Author(s):

W. R. Winnall ◽

J. Gould ◽

J. A. Muir ◽

P. Hertzog ◽

M. P. Hedger

Keyword(s):

Gene Expression ◽

Microarray Analysis ◽

Inflammatory Mediators ◽

Inflammatory Cytokine ◽

Transforming Growth Factor ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Gene Expression Response ◽

Anti Inflammatory ◽

Testicular Macrophages

Studies on rat testicular macrophages (TMs) have indicated that these cells play an important role in testis function by supporting the immunosuppressive environment that protects developing germ cells and by responding to pathogens. By comparison, mouse TMs are essentially uncharacterised due to difficulties in isolating sufficient cells for study. We have established a technique for isolating 95% pure TMs from adult mice by differential adherence. Mouse TMs were cultured for 3h with saline, 10 or 100 ng/mL lipopolysaccharide (LPS) and compared with resident peritoneal macrophages (PMs) and bone marrow-derived macrophages (BMMs). Expression of inflammatory regulators was determined using real-time Q-PCR and AgilentTM microarray analysis. Microarray analysis indicated that each macrophage type displayed very distinct gene expression profiles. There were 526 genes uniquely expressed in TMs at basal levels compared with the other macrophages and 268 genes uniquely expressed by TMs after LPS treatment. Q-PCR determined that LPS induced expression of the anti-inflammatory cytokine interleukin (IL)-10 in each of the macrophage types, with BMMs the strongest responders. LPS stimulated IL-10 mRNA approximately 100-fold in TMs, but only 20-fold in PMs. The anti-inflammatory transforming growth factor-β1 was not significantly induced at this time-point in any macrophage type. In terms of pro-inflammatory mediators, the TM response to LPS was always lower compared to the BMMs. Compared to PMs, the responses of TMs were similar for the hallmark pro-inflammatory cytokine tumour necrosis factor- a, but 40% less for IL-1β. TMs were also deficient in production of IL-6 and cyclooxygenase-2 and IL-12. TMs were therefore relatively strong responders to LPS in terms of IL-10, but weak responders in terms of pro-inflammatory mediators, indicating an immunosuppressive phenotype. The isolation and gene measurement methods established in this study will allow us to use knockout and transgenic mouse models to determine the role for TMs in testicular inflammation/fibrosis models.

Download Full-text

Timing of gene expression and recruitment in independent origins of CAM in the Agavoideae (Asparagaceae).

10.1101/2021.11.10.468112 ◽

2021 ◽

Author(s):

Karolina Heyduk ◽

Edward McAssey ◽

James Leebens-Mack

Keyword(s):

Gene Expression ◽

Carbon Fixation ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Transcriptional Response ◽

Time Of Day ◽

Temporal Separation ◽

Genes Encoding ◽

Cam Species ◽

Cam Photosynthesis

CAM photosynthesis has evolved repeatedly across the plant tree of life, yet our understanding of the genetic convergence across independent origins remains hampered by the lack of comparative studies. CAM is furthermore thought to be closely linked to the circadian clock in order to achieve temporal separation of carboxylation and sugar production. Here, we explore gene expression profiles in eight species from the Agavoideae (Asparagaceae) encompassing three independent origins of CAM. Using comparative physiology and transcriptomics, we examined the variable modes of CAM in this subfamily and the changes in gene expression across time of day and between well-watered and drought-stressed treatments. We further assessed gene expression and molecular evolution of genes encoding phosphoenolpyruvate carboxylase (PPC), an enzyme required for primary carbon fixation in CAM. Most time-of-day expression profiles are largely conserved across all eight species and suggest that large perturbations to the central clock are not required for CAM evolution. In contrast, transcriptional response to drought is highly lineage specific. Yucca and Beschorneria have CAM-like expression of PPC2, a copy of PPC that has never been shown to be recruited for CAM in angiosperms, and evidence of positive selection in PPC genes implicates mutations that may have facilitated the recruitment for CAM function early in the evolutionary history of the Agavoideae. Together the physiological and transcriptomic comparison of closely related C3 and CAM species reveals similar gene expression profiles, with the notable exception of differential recruitment of carboxylase enzymes for CAM function.

Download Full-text

Transcripts from downstream alternative transcription start sites evade uORF-mediated inhibition of gene expression in Arabidopsis

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1804971115 ◽

2018 ◽

Vol 115 (30) ◽

pp. 7831-7836 ◽

Cited By ~ 29

Author(s):

Yukio Kurihara ◽

Yuko Makita ◽

Mika Kawashima ◽

Tomoya Fujita ◽

Shintaro Iwasaki ◽

...

Keyword(s):

Gene Expression ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Transcript Abundance ◽

Initial Growth ◽

Transcription Start ◽

Transcription Start Sites ◽

Genome Wide ◽

Nonsense Mediated Mrna Decay ◽

Inducible Genes

Plants adapt to alterations in light conditions by controlling their gene expression profiles. Expression of light-inducible genes is transcriptionally induced by transcription factors such as HY5. However, few detailed analyses have been carried out on the control of transcription start sites (TSSs). Of the various wavelengths of light, it is blue light (BL) that regulates physiological responses such as hypocotyl elongation and flowering time. To understand how gene expression is controlled not only by transcript abundance but also by TSS selection, we examined genome-wide TSS profiles in Arabidopsis seedlings after exposure to BL irradiation following initial growth in the dark. Thousands of genes use multiple TSSs, and some transcripts have upstream ORFs (uORFs) that take precedence over the main ORF (mORF) encoding proteins. The uORFs often function as translation inhibitors of the mORF or as triggers of nonsense-mediated mRNA decay (NMD). Transcription from TSSs located downstream of the uORFs in 220 genes is enhanced by BL exposure. This type of regulation is found in HY5 and HYH, major regulators of light-dependent gene expression. Translation efficiencies of the genes showing enhanced usage of these TSSs increased upon BL exposure. We also show that transcripts from TSSs upstream of uORFs in 45 of the 220 genes, including HY5, accumulated in a mutant of NMD. These results suggest that BL controls gene expression not only by enhancing transcriptions but also by choosing the TSS, and transcripts from downstream TSSs evade uORF-mediated inhibition to ensure high expression of light-regulated genes.

Download Full-text

Validating the Cyc2 neutrophilic Fe oxidation pathway using meta-omics of Zetaproteobacteria iron mats at marine hydrothermal vents

10.1101/722066 ◽

2019 ◽

Cited By ~ 6

Author(s):

Sean M. McAllister ◽

Shawn W. Polson ◽

David A. Butterfield ◽

Brian T. Glazer ◽

Jason B. Sylvan ◽

...

Keyword(s):

Hydrothermal Vents ◽

Carbon Fixation ◽

Central Metabolism ◽

Ideal Model ◽

Oxidase Gene ◽

Oxidation Pathway ◽

Fe Oxidation ◽

Mid Atlantic Ridge ◽

Metabolism Gene

AbstractZetaproteobacteria create extensive iron (Fe) oxide mats at marine hydrothermal vents, making them an ideal model for microbial Fe oxidation at circumneutral pH. Comparison of neutrophilic Fe-oxidizer isolate genomes has revealed a hypothetical Fe oxidation pathway, featuring a homolog of the Fe oxidase Cyc2 from Acidithiobacillus ferrooxidans. However, Cyc2 function is not well verified in neutrophilic Fe-oxidizers, particularly in Fe-oxidizing environments. Toward this, we analyzed genomes and metatranscriptomes of Zetaproteobacteria, using 53 new high-quality metagenome assembled genomes reconstructed from Fe mats at Mid-Atlantic Ridge, Mariana Backarc, and Loihi Seamount (Hawaii) hydrothermal vents. Phylogenetic analysis demonstrated conservation of Cyc2 sequences among most neutrophilic Fe-oxidizers, suggesting a common function. We confirmed the widespread distribution of cyc2 and other model Fe oxidation pathway genes across all represented Zetaproteobacteria lineages. High expression of these genes was observed in diverse Zetaproteobacteria under multiple environmental conditions, and in incubations. The putative Fe oxidase gene, cyc2, was highly expressed in situ, often as the top expressed gene. The cyc2 gene showed increased expression in Fe(II)-amended incubations, with corresponding increases in carbon fixation and central metabolism gene expression. These results substantiate the Cyc2-based Fe oxidation pathway in neutrophiles and demonstrate its significance in marine Fe-mineralizing environments.

Download Full-text

LBH589, a novel histone deacetylase inhibitor (HDACi), treatment of patients with cutaneous T-cell lymphoma (CTCL). Changes in skin gene expression profiles related to clinical response following therapy

Journal of Clinical Oncology ◽

10.1200/jco.2006.24.18_suppl.7501 ◽

2006 ◽

Vol 24 (18_suppl) ◽

pp. 7501-7501 ◽

Cited By ~ 5

Author(s):

H. M. Prince ◽

D. J. George ◽

R. Johnstone ◽

R. Williams-Truax ◽

P. Atadja ◽

...

Keyword(s):

Gene Expression ◽

Histone Deacetylase ◽

Dose Level ◽

Histone Deacetylase Inhibitor ◽

Time Course ◽

Expression Profiles ◽

Gene Expression Response ◽

Deacetylase Inhibitor ◽

Grade Iii

7501 Background: LBH589 is a novel histone deacetylase inhibitor in Phase I trials. Since other HDACi have induced disease regression in CTCL, we evaluated the activity of LBH589 and resulting changes in gene expression of LBH589 in CTCL patients (pts). Methods: Pts with advanced-stage CTCL, who had progressed following prior systemic therapy were entered into the DLT dose level 30mg M,W,F cohort (n = 1) or the subsequent MTD dose level 20mg M,W, F weekly (n = 10). LBH589 was continued until disease progression or unacceptable toxicity. The first three pts had 3mm punch biopsies from CTCL-involved skin lesions at 0, 4, 8 and 24h after administration, which were subjected to gene expression profiling using Affymetrix U133 plus 2.0 GeneChips with 47,000 probesets. Results: Eleven pts with CTCL have been entered to date. Two of the pts attained a complete response (CR), 3 attained a partial response (PR), 2 achieved stable disease (SD) with ongoing improvement, and 4 progressed on treatment (PD). Of particular interest, 2 pts who were initially SD required discontinuation because of toxicities (Grade III diarrhea at week 4, Grade II fatigue at week 12). Both had ongoing improvement in their disease achieving a CR and PR, respectively 3 months later. Of the 5 responding pts, one with a CR (discontinued after 10 doses due to Grade III diarrhea) progressed at 8m. Microarray data on the first 3 pts (2CR and 1PD) demonstrated distinct gene expression response profiles between the 3 pts. Surprisingly, the pt with PD showed the greatest transcriptional response with more than 16,000 genes activated or repressed over the 24 hr time course. Of these responsive genes, close to 60% were activated while 40% were repressed. In contrast, less than 1000 genes showed a 2-fold change in expression in the 2 pts with a CR with greater than 85% of the genes being repressed. Conclusions: LBH589 induce CR’s in CTCL pts. Regression of disease can occur some weeks after discontinuation of therapy. Preliminary microarray analysis of tumor samples indicated that LBH589 mediates changes in gene expression in vivo with an unexpected observed inverse relationship between the number of genes altered and clinical outcome. [Table: see text]

Download Full-text