Project description:The potency and indiscriminate nature of formaldehyde reactivity upon biological molecules make it a universal stressor. However, some organisms such as Methylorubrum extorquens possess means to rapidly and effectively mitigate formaldehyde-induced damage. EfgA is a recently identified formaldehyde sensor predicted to halt translation in response to elevated formaldehyde as a means to protect cells. Herein, we investigate growth, formaldehyde consumption, and changes in gene expression to better understand how M. extorquens responds to formaldehyde with and without the EfgA-formaldehyde-mediated translational response, and how this mechanism compares to other forms of translation inhibition. These distinct mechanisms of translation inhibition have notable differences: they each involve different specific players and in addition, formaldehyde also acts as a general, multi-target stressor and a potential carbon source. Here, we present findings demonstrating that in addition to its characterized impact on translation, functional EfgA also allows for a rapid and robust transcriptional response to formaldehyde and that removal of efgA leads to heightened proteotoxic and genotoxic stress in the presence of increased formaldehyde levels. As our previous work suggested that formaldehyde is proteotoxic in M. extorquens, we accurately predicted that strains lacking efgA would experience increased protein damage. We also found that many downstream consequences of translation inhibition were shared by EfgA-formaldehyde- and kanamycin-mediated translation inhibition. Our work to modularize the transcriptional response uncovered additional layers of regulatory control enacted by functional EfgA upon experiencing formaldehyde stress, and further demonstrate the importance this protein plays at both transcriptional and translational levels in this model methylotroph.
Project description:We are investigating the miRNA expression profiles of human lung cells to gaseous formaldehyde We used microarrays to detail the global programme of miRNA expression upon response to formaldehyde
Project description:Formaldehyde is a toxic volatile organic compound and its mechanism of toxicity to plant has not yet been revealed. This experiment was designed to identify formaldehyde-responsible genes under exposure to low or high concentration of airborne formaldehyde for a short period of time. 7 weeks old Arabidopsis thaliana transformant (ecotype: Columbia) plants were exposed to gaseous formaldehyde at 1-2 ppm (low), 14-16 ppm (high), or less than 0.04 ppm (air control) at 24oC under light-condition for 150 minutes inside a chamber for formaldehyde exposure. Total RNA was isolated from rosette leaves of exposed plants and was applied to microarray analysis. We investigated into formaldehyde dose response on gene expression of Arabidopsis and tried to understand the toxic mechanisms of formaldehyde using an Affymetrix Arabidopsis genome array ATH-1.
Project description:A number of reports showed that photo-bio-modulation by various lasers has favorable biological effects on cells. However, the effects of low-level Er:YAG laser irradiation on osteoblasts remain unclear. The purpose of this study was to evaluate the effects on proliferation and osteogenic differentiation of primary osteoblast-like cells by low-level Er:YAG laser irradiation. Osteoblast-like cells isolated from the calvariae of 3–5-day-old Wistar rats were irradiated by Er:YAG laser at energy fluences of 2.2, 3.3, or 4.3 J/cm2, respectively. After irradiation, cell surface temperature was measured and cell proliferation was evaluated by flow cytometry. Calcification was evaluated by measuring the Alizarin red S staining area after 7-day-culture with osteoinductive medium. Gene expressions in non-irradiated and laser-irradiated cells were evaluated by qPCR at 3, 6, and 12 hours after irradiation. Microarray analysis was performed to comprehensively evaluate gene expressions of non-irradiated cells and irradiated cells at 3.3 J/cm2 at 6 hours after irradiation. No pronounced increase of cell surface temperature was induced by low-level Er:YAG laser irradiation, and the Er:YAG laser irradiation did not affect osteoblast-like cell proliferation. Osteoblast-like cell calcification was significantly increased 7 days after Er:YAG laser irradiation at 3.3 J/cm2. Bglap expression was significantly increased in cells irradiated at 3.3 J/cm2 at 6 hours post-irradiation. Microarray analysis showed that irradiation at 3.3 J/cm2 caused up-regulation of inflammation-related genes and down-regulation of Wisp2. Gene set enrichment analysis also clarified enrichment of inflammation-related gene sets and Notch signaling pathway. In conclusion, Low-level Er:YAG laser irradiation enhanced calcification of primary osteoblast-like cells via enhanced Bglap expression and enriched Notch signaling pathway.
Project description:In order for bacteria to thrive, they must be well-adapted to their environmental niche, which may involve specialized metabolism, timely adaptation to shifting environments, and/or the ability to mitigate numerous stressors. These attributes are highly dependent on cellular machinery that can sense both the external and intracellular environment. Methylorubrum extorquens is an extensively studied facultative methylotroph, an organism that can use single-carbon compounds as their sole source of carbon and energy. In methylotrophic metabolism, carbon flows through formaldehyde as a central metabolite; thus, formaldehyde is both an obligate metabolite and a metabolic stressor. Via the one-carbon dissimilation pathway, free formaldehyde is rapidly incorporated by formaldehyde activating enzyme (Fae), which is constitutively expressed at high levels. In the presence of elevated formaldehyde levels, a recently identified formaldehyde-sensing protein, EfgA, induces growth arrest. Herein, we describe TtmR, a formaldehyde-responsive transcription factor that, like EfgA, modulates formaldehyde resistance. It is a member of the MarR family of transcription factors and impacts the expression of 75 genes distributed throughout the genome, many of which are themselves transcription factors and/or involved in stress response, including efgA. Notably, when M. extorquens is adapting its metabolic network during the transition to methylotrophy, efgA and ttmR mutants experience an imbalance in formaldehyde production and a notable growth delay. Although methylotrophy necessitates that M. extorquens maintain a relatively high level of formaldehyde tolerance, this work reveals a tradeoff between formaldehyde resistance and the efficient transition to methylotrophic growth and suggests that TtmR and EfgA play a pivotal role in maintaining this balance.
Project description:In addition to gaining knowledge on in vivo miRNA responses to formaldehyde, we set out to relate these miRNA responses to transcriptional profiles modified by formaldehyde. Rats were exposed by inhalation to either 0 or 2 ppm formaldehyde (6 hours/day) for 28 days. Genome-wide transcriptional profiles and associated signaling pathways were assessed within the nasal respiratory mucosa and circulating mononuclear white blood cells (WBC).
Project description:Reactive aldehydes are produced during cellular metabolism and can accumulate in specific tissues particularly when aldehyde clearance mechanisms are impaired. Reactive aldehydes induce DNA-DNA and DNA-protein crosslinks (DPCs) that are repaired by different DNA repair pathways. Cellular toxicity of endogenous formaldehyde has been attributed to the damage of the genomic DNA and consequent inhibition of transcription. However, whether damage to other cellular macromolecules and interference with additional metabolic processes contributes to formaldehyde toxicity has not been investigated. We demonstrate that formaldehyde induces toxic RNA-protein crosslinks (RPCs) in mRNA that inhibit translation and induce a specific RPC stress response pathway that is characterized by linkage-specific ubiquitylation. RPCs in the mRNA are recognized by stalling ribosomes and marked by K6-linked ubiquitylation that promotes their clearance by the ubiquitin-dependent unfoldase VCP.
Project description:One-carbon metabolism is an essential branch of cellular metabolism that intersects with epigenetic regulation. Here, we show formaldehyde, a one-carbon unit derived from both endogenous sources and environmental exposure, regulates one-carbon metabolism by inhibiting the biosynthesis of S-adenosylmethionine (SAM), the major methyl donor in cells. Formaldehyde reacts with privileged, hyperreactive cysteine sites in the proteome, including Cys120 in S-adenosylmethionine synthase isoform type-1 (MAT1A). Formaldehyde exposure inhibited MAT1A activity and decreased SAM production with MAT-isoform specificity. A genetic mouse model of chronic formaldehyde overload showed a decrease in SAM and in methylation on selected histones and genes. Epigenetic and transcriptional regulation of Mat1a and related genes function as compensatory mechanisms for formaldehyde-dependent SAM depletion, revealing a biochemical feedback cycle between formaldehyde and SAM one-carbon units.
Project description:We set out to test the hypothesis that formaldehyde inhalation exposure significantly alters miRNA expression profiles within the nasal epithelium of nonhuman primates. Here, cynomolgus macaques were exposed to 0, 2, and 6 ppm formaldehyde for 6 hours/day across two consecutive days. RNA was extracted from the nasal maxilloturbinate region, a direct target of formaldehyde inhalation exposure. Genome-wide miRNA expression levels were assessed using microarrays.