Free spermidine evokes superoxide radicals that manifest toxicity
- CSIR Institute of Microbial Technology, India
- St John's Medical College Hospital, India
Abstract
Spermidine and other polyamines alleviate oxidative stress, yet excess spermidine seems toxic to Escherichia coli unless it is neutralized by SpeG, an enzyme for the spermidine N-acetyl transferase function. Thus, wild-type E. coli can tolerate applied exogenous spermidine stress, but DspeG strain of E. coli fails to do that. Here, using different ROS probes and performing electron paramagnetic resonance spectroscopy, we provide evidence that although spermidine mitigates oxidative stress by lowering overall ROS levels, excess of it simultaneously triggers the production of superoxide radicals, thereby causing toxicity in the DspeG strain. Furthermore, performing microarray experiment and other biochemical assays, we show that the spermidine-induced superoxide anions affected redox balance and iron homeostasis. Finally, we demonstrate that while RNA-bound spermidine inhibits iron oxidation, free spermidine interacts and oxidizes the iron to evoke superoxide radicals directly. Therefore, we propose that the spermidine-induced superoxide generation is one of the major causes of spermidine toxicity in E. coli.
Data availability
Microarray data is available in the GEO server. GEO accession Number GSE154618 has been provided in the material and method section.Source files for the following Figures were provided as a zip folder:Figure 1A, 1B, 1C, 1FFigure 2Figure 3A, 3B, 3C, 3D, 3E, 3F, 3GFigure 4B (ii), 4C, 4D, 4EFigure 5A, 5B, 5DFigure 6D, 6E, 6GFigure 1-figure supplement 1C
-
The global transcriptomic profile in the spermidine-stressed E. coliNCBI Gene Expression Omnibus, GSE154618.
Article and author information
Author details
Funding
Council of Scientific and Industrial Research, India (MLP042)
- Dipak Dutta
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Joseph T Wade, New York State Department of Health, United States
Version history
- Preprint posted: September 5, 2021 (view preprint)
- Received: February 8, 2022
- Accepted: April 11, 2022
- Accepted Manuscript published: April 13, 2022 (version 1)
- Accepted Manuscript updated: April 14, 2022 (version 2)
- Version of Record published: April 25, 2022 (version 3)
Copyright
© 2022, Kumar et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Metrics
-
- 2,332
- views
-
- 295
- downloads
-
- 13
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Download links
A two-part list of links to download the article, or parts of the article, in various formats.
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Free spermidine evokes superoxide radicals that manifest toxicity
eLife 11:e77704.
https://doi.org/10.7554/eLife.77704
Further reading
-
- Biochemistry and Chemical Biology
- Neuroscience
Amyloid β (Aβ) peptides accumulating in the brain are proposed to trigger Alzheimer’s disease (AD). However, molecular cascades underlying their toxicity are poorly defined. Here, we explored a novel hypothesis for Aβ42 toxicity that arises from its proven affinity for γ-secretases. We hypothesized that the reported increases in Aβ42, particularly in the endolysosomal compartment, promote the establishment of a product feedback inhibitory mechanism on γ-secretases, and thereby impair downstream signaling events. We conducted kinetic analyses of γ-secretase activity in cell-free systems in the presence of Aβ, as well as cell-based and ex vivo assays in neuronal cell lines, neurons, and brain synaptosomes to assess the impact of Aβ on γ-secretases. We show that human Aβ42 peptides, but neither murine Aβ42 nor human Aβ17–42 (p3), inhibit γ-secretases and trigger accumulation of unprocessed substrates in neurons, including C-terminal fragments (CTFs) of APP, p75, and pan-cadherin. Moreover, Aβ42 treatment dysregulated cellular homeostasis, as shown by the induction of p75-dependent neuronal death in two distinct cellular systems. Our findings raise the possibility that pathological elevations in Aβ42 contribute to cellular toxicity via the γ-secretase inhibition, and provide a novel conceptual framework to address Aβ toxicity in the context of γ-secretase-dependent homeostatic signaling.
-
- Biochemistry and Chemical Biology
- Cell Biology
Protein homeostasis (proteostasis) deficiency is an important contributing factor to neurological and metabolic diseases. However, how the proteostasis network orchestrates the folding and assembly of multi-subunit membrane proteins is poorly understood. Previous proteomics studies identified Hsp47 (Gene: SERPINH1), a heat shock protein in the endoplasmic reticulum lumen, as the most enriched interacting chaperone for gamma-aminobutyric acid type A (GABAA) receptors. Here, we show that Hsp47 enhances the functional surface expression of GABAA receptors in rat neurons and human HEK293T cells. Furthermore, molecular mechanism study demonstrates that Hsp47 acts after BiP (Gene: HSPA5) and preferentially binds the folded conformation of GABAA receptors without inducing the unfolded protein response in HEK293T cells. Therefore, Hsp47 promotes the subunit-subunit interaction, the receptor assembly process, and the anterograde trafficking of GABAA receptors. Overexpressing Hsp47 is sufficient to correct the surface expression and function of epilepsy-associated GABAA receptor variants in HEK293T cells. Hsp47 also promotes the surface trafficking of other Cys-loop receptors, including nicotinic acetylcholine receptors and serotonin type 3 receptors in HEK293T cells. Therefore, in addition to its known function as a collagen chaperone, this work establishes that Hsp47 plays a critical and general role in the maturation of multi-subunit Cys-loop neuroreceptors.
