An FYVE-Domain-Containing Protein, PsFP1, Is Involved in Vegetative Growth, Oxidative Stress Response and Virulence of Phytophthora sojae

Proteins that contain the FYVE zinc-finger domain are recruited to PtdIns3P-containing membranes, participating in numerous biological processes such as membrane trafficking, cytoskeletal regulation, and receptor signaling. However, the genome-wide distribution, evolution, and biological functions of FYVE-containing proteins are rarely reported for oomycetes. By genome mining of Phytophthora sojae, two proteins (PsFP1 and PsFP2) with a combination of the FYVE domain and the PX domain (a major phosphoinositide binding module) were found. To clarify the functions of PsFP1 and PsFP2, the CRISPR/Cas9-mediated gene replacement system was used to knock out the two genes respectively. Only heterozygous deletion mutants of PsFP1 were recovered, and the expression level of PsFP1 in the heterozygous knockout transformants was significantly down-regulated. These PsFP1 mutants showed a decrease in mycelial growth and pathogenicity and were more sensitive to hydrogen peroxide. These phenotypes were recovered to the level of wild-type by overexpression PsFP1 gene in the PsFP1 heterozygous knockout transformant. In contrast, deletion of PsFP2 had no significant effect on vegetative growth, asexual and sexual reproduction, pathogenicity, or oxidative stress sensitivity. PsFP1 was primarily localized in vesicle-like structures and both the FYVE and PX domains are important for its localization. Overall, our results indicate that PsFP1 plays an important role in the vegetative growth and virulence of P. sojae.

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NAD(P)H Cytochrome b5 Oxidoreductase Deficiency in Leishmania major Results in Impaired Linoleate Synthesis Followed by Increased Oxidative Stress and Cell Death

Journal of Biological Chemistry ◽

10.1074/jbc.m112.389338 ◽

2012 ◽

Vol 287 (42) ◽

pp. 34992-35003 ◽

Cited By ~ 13

Author(s):

Supratim Mukherjee ◽

Sumit Sen Santara ◽

Shantanabha Das ◽

Moumita Bose ◽

Jayasree Roy ◽

...

Keyword(s):

Oxidative Stress ◽

Cell Death ◽

Leishmania Major ◽

Flow Cytometric Analysis ◽

Physiological Role ◽

Gene Replacement ◽

Spectral Studies ◽

Null Cell ◽

Knock Out ◽

Δ9 Desaturase

NAD(P)H cytochrome b5 oxidoreductase (Ncb5or), comprising cytochrome b5 and cytochrome b5 reductase domains, is widely distributed in eukaryotic organisms. Although Ncb5or plays a crucial role in lipid metabolism of mice, so far no Ncb5or gene has been reported in the unicellular parasitic protozoa Leishmania species. We have cloned, expressed, and characterized Ncb5or gene from Leishmania major. Steady state catalysis and spectral studies show that NADH can quickly reduce the ferric state of the enzyme to the ferrous state and is able to donate an electron(s) to external acceptors. To elucidate its exact physiological role in Leishmania, we attempted to create NAD(P)H cytochrome b5 oxidoreductase from L. major (LmNcb5or) knock-out mutants by targeted gene replacement technique. A free fatty acid profile in knock-out (KO) cells reveals marked deficiency in linoleate and linolenate when compared with wild type (WT) or overexpressing cells. KO culture has a higher percentage of dead cells compared with both WT and overexpressing cells. Increased O2 uptake, uncoupling and ATP synthesis, and loss of mitochondrial membrane potential are evident in KO cells. Flow cytometric analysis reveals the presence of a higher concentration of intracellular H2O2, indicative of increased oxidative stress in parasites lacking LmNcb5or. Cell death is significantly reduced when the KO cells are pretreated with BSA bound linoleate. Real time PCR studies demonstrate a higher Δ12 desaturase, superoxide dismutase, and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) mRNA with a concomitant fall in Δ9 desaturase mRNA expression in LmNcb5or null cell line. Together these findings suggest that decreased linoleate synthesis, and increased oxidative stress and apoptosis are the major consequences of LmNcb5or deficiency in Leishmania.

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Participation of Rho-dependent transcription termination in oxidative stress sensitivity caused by an rpoB mutation

Genes to Cells ◽

10.1111/j.1365-2443.2005.00849.x ◽

2005 ◽

Vol 10 (5) ◽

pp. 477-487 ◽

Cited By ~ 9

Author(s):

Nobuyuki Kawamura ◽

Kenji Kurokawa ◽

Takahiro Ito ◽

Hiroshi Hamamoto ◽

Hiroshi Koyama ◽

...

Keyword(s):

Oxidative Stress ◽

Transcription Termination ◽

Stress Sensitivity ◽

Rpob Mutation

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Primed to die: an investigation of the genetic mechanisms underlying noise-induced hearing loss and cochlear damage in homozygous Foxo3-knockout mice

Cell Death and Disease ◽

10.1038/s41419-021-03972-6 ◽

2021 ◽

Vol 12 (7) ◽

Author(s):

Holly J. Beaulac ◽

Felicia Gilels ◽

Jingyuan Zhang ◽

Sarah Jeoung ◽

Patricia M. White

Keyword(s):

Oxidative Stress ◽

Hearing Loss ◽

Noise Exposure ◽

Multiphoton Microscopy ◽

Cell Loss ◽

Outer Hair Cells ◽

Noise Induced Hearing Loss ◽

Knock Out ◽

Cell Death Pathway ◽

Cochlear Damage

AbstractThe prevalence of noise-induced hearing loss (NIHL) continues to increase, with limited therapies available for individuals with cochlear damage. We have previously established that the transcription factor FOXO3 is necessary to preserve outer hair cells (OHCs) and hearing thresholds up to two weeks following mild noise exposure in mice. The mechanisms by which FOXO3 preserves cochlear cells and function are unknown. In this study, we analyzed the immediate effects of mild noise exposure on wild-type, Foxo3 heterozygous (Foxo3+/−), and Foxo3 knock-out (Foxo3−/−) mice to better understand FOXO3’s role(s) in the mammalian cochlea. We used confocal and multiphoton microscopy to examine well-characterized components of noise-induced damage including calcium regulators, oxidative stress, necrosis, and caspase-dependent and caspase-independent apoptosis. Lower immunoreactivity of the calcium buffer Oncomodulin in Foxo3−/− OHCs correlated with cell loss beginning 4 h post-noise exposure. Using immunohistochemistry, we identified parthanatos as the cell death pathway for OHCs. Oxidative stress response pathways were not significantly altered in FOXO3’s absence. We used RNA sequencing to identify and RT-qPCR to confirm differentially expressed genes. We further investigated a gene downregulated in the unexposed Foxo3−/− mice that may contribute to OHC noise susceptibility. Glycerophosphodiester phosphodiesterase domain containing 3 (GDPD3), a possible endogenous source of lysophosphatidic acid (LPA), has not previously been described in the cochlea. As LPA reduces OHC loss after severe noise exposure, we treated noise-exposed Foxo3−/− mice with exogenous LPA. LPA treatment delayed immediate damage to OHCs but was insufficient to ultimately prevent their death or prevent hearing loss. These results suggest that FOXO3 acts prior to acoustic insult to maintain cochlear resilience, possibly through sustaining endogenous LPA levels.

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Acrolein-Induced Oxidative Stress In NAD(P)H Oxidase Subunit gp91phox Knock-Out Mice and its modulation of NFκB and CD36

Journal of Health Care for the Poor and Underserved ◽

10.1353/hpu.2014.0002 ◽

2014 ◽

Vol 24 (4A) ◽

pp. 118-131 ◽

Cited By ~ 4

Author(s):

Zivar Yousefipour ◽

Chelsea Zhang ◽

Mahdieh Monfareed ◽

James Walker ◽

Mohammad Newaz

Keyword(s):

Oxidative Stress ◽

Knock Out ◽

Oxidase Subunit ◽

Knock Out Mice

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Abstract 16: Progressive Alterations Of Mitochondrial Bioenergetics In The Kidney During The Development Of Salt-induced Hypertension

Hypertension ◽

10.1161/hyp.76.suppl_1.16 ◽

2020 ◽

Vol 76 (Suppl_1) ◽

Author(s):

Namrata Tomar ◽

Sunil M Kandel ◽

Xiao Zhang ◽

Nadezhda Zheleznova ◽

Allen W Cowley ◽

...

Keyword(s):

Oxidative Stress ◽

Complex Disease ◽

Atp Synthesis ◽

Control Group ◽

Mitochondrial Uncoupling ◽

Knock Out ◽

Atp Production ◽

Consumption Rates ◽

Mitochondrial Efficiency ◽

Salt Sensitive Hypertension

Hypertension is a complex disease and a leading cause of morbidity and mortality globally. Although oxidative stress and mitochondrial dysfunction have been found in the kidney in various models of hypertension, progressive alteration of mitochondrial oxidative phosphorylation (OxPhos) in the kidney during the development of salt-sensitive hypertension has not been characterized. The present study determined changes of OxPhos in kidneys of Dahl salt-sensitive (SS) rats before (0.4% NaCl diet; LS) and after switching to a high salt diet (4.0% NaCl; HS) during the development of hypertension. Mitochondria were isolated from the outer medulla (OM) and cortex of the kidney of SS rats fed a LS diet since weaning and studied at days 3, 7, 14 & 21 of a HS diet feeding. Oxygen consumption rates (OCR) were measured in mitochondria energized with pyruvate + malate as substrates for three different respiratory states using an Oroboros Oxygraph-2k Instrument. This includes i) leak state (in the absence of ADP), ii) ADP-stimulated state, and iii) uncoupled state (in the presence of an uncoupler FCCP). A biphasic pattern of ADP-stimulated OCR with progressive uncoupling was observed in both the renal OM and cortex. Mitochondrial efficiency for ATP synthesis was increased in the early phases of hypertension (3 & 7 days) but was severely compromised in the established phases of hypertension (14 & 21 days). This decreased mitochondrial efficiency was associated with uncoupling of OxPhos and high levels of oxidative stress which we hypothesized were due to mitochondrial ROS stimulation of membrane NOXs. To test this, experiments were performed in SS rats with double knock out (DKO) of the cytosolic subunit of NOX2 (p67 phox ) and NOX4 (SS p67phox-/-/Nox4-/- ). DKO SS rats were fed a HS diet and OCR of renal cortical and OM mitochondria was determined at days 7 and 14. In contrast to SS rats, the DKO SS rats fed a HS diet showed no significant differences in mitochondrial OCR in the cortex or OM, nor to a control group maintained on a LS diet. HS diet in SS rats initially increases the efficiency of renal cortical and medullary mitochondrial ATP production (days 1-7) followed by an enhanced ROS production with mitochondrial uncoupling and reduced efficiency of ATP production by the third week.

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Abstract MP124: De-palmitoylation of Cysteine 202 of Cyclophilin-D by Calcium is Important for the Activation of the Permeability Transition Pore

Circulation Research ◽

10.1161/res.127.suppl_1.mp124 ◽

2020 ◽

Vol 127 (Suppl_1) ◽

Author(s):

Georgios Amanakis ◽

Junhui Sun ◽

Maria Fergusson ◽

Chengyu Liu ◽

Jeff D Molkentin ◽

...

Keyword(s):

Oxidative Stress ◽

Mitochondrial Permeability Transition ◽

Ischemia Reperfusion ◽

Permeability Transition Pore ◽

Permeability Transition ◽

Calcium Overload ◽

Mitochondrial Calcium ◽

Cyclophilin D ◽

Perfused Heart ◽

Knock Out

Cyclophilin-D (CypD) is a well-known regulator of the mitochondrial permeability transition pore (PTP), the main effector of cardiac ischemia/reperfusion (I/R) injury characterized by oxidative stress and calcium overload. However, the mechanism by which CypD activates PTP is poorly understood. Cysteine 202 of CypD (C202) is highly conserved across species and can undergo redox-sensitive post-translational modifications, such as S-nitrosylation and oxidation. To study the importance of C202, we developed a knock-in mouse model using CRISPR where CypD-C202 was mutated to a serine (C202S). Hearts from these mice are protected against I/R injury. We found C202 to be abundantly S-palmitoylated under baseline conditions while C202 was de-palmitoylated during ischemia in WT hearts. To further investigate the mechanism of de-palmitoylation during ischemia, we considered the increase of matrix calcium, oxidative stress and uncoupling of ATP synthesis from the electron transport chain. We tested the effects of these conditions on the palmitoylation of CypD in isolated cardiac mitochondria. The palmitoylation of CypD was assessed using a resin-assisted capture (Acyl-RAC). We report that oxidative stress (phenylarsenide) and uncoupling (CCCP) had no effect on CypD palmitoylation (p>0.05, n=3 and n=7 respectively). However, calcium overload led to de-palmitoylation of CypD to the level observed at the end ischemia (1±0.10 vs 0.63±0.09, p=0.012, n=9). To further test the hypothesis that calcium regulates S-palmitoylation of CypD we measured S-palmitoylation of CypD in non-perfused heart lysates from global germline mitochondrial calcium uniporter knock-out mice (MCU-KO), which have reduced mitochondrial calcium and we found an increase in S-palmitoylation of CypD (WT 1±0.04 vs MCU-KO 1.603±0.11, p<0.001, n=6). The data are consistent with the hypothesis that C202 is important for the CypD mediated activation of PTP. Ischemia leads to increased matrix calcium which in turn promotes the de-palmitoylation of CypD on C202. The now free C202 can further be oxidized during reperfusion leading to the activation of PTP. Thus, S-palmitoylation and oxidation of CypD-C202 possibly target CypD to the PTP, making them potent regulators of cardiac I/R injury.

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Transcriptome analysis of LRRK2 knock-out microglia cells reveals alterations of inflammatory- and oxidative stress-related pathways upon treatment with α-synuclein fibrils

Neurobiology of Disease ◽

10.1016/j.nbd.2019.05.012 ◽

2019 ◽

Vol 129 ◽

pp. 67-78 ◽

Cited By ~ 16

Author(s):

Isabella Russo ◽

Alice Kaganovich ◽

Jinhui Ding ◽

Natalie Landeck ◽

Adamantios Mamais ◽

...

Keyword(s):

Oxidative Stress ◽

Transcriptome Analysis ◽

Knock Out ◽

And Oxidative Stress

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Oxidative stress sensitivity of engineered Escherichia coli cells with a reduced genome

FEMS Microbiology Letters ◽

10.1111/j.1574-6968.2011.02331.x ◽

2011 ◽

Vol 322 (1) ◽

pp. 25-33 ◽

Cited By ~ 23

Author(s):

Yumi Iwadate ◽

Hirofumi Honda ◽

Haruhiko Sato ◽

Masayuki Hashimoto ◽

Jun-ichi Kato

Keyword(s):

Oxidative Stress ◽

Escherichia Coli ◽

Stress Sensitivity ◽

Reduced Genome ◽

Escherichia Coli Cells ◽

Engineered Escherichia Coli

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Cyanamide Phytotoxicity in Soybean (Glycine max) Seedlings involves Aldehyde Dehydrogenase Inhibition and Oxidative Stress

Natural Product Communications ◽

10.1177/1934578x1501000511 ◽

2015 ◽

Vol 10 (5) ◽

pp. 1934578X1501000

Author(s):

John S. Maninang ◽

Shin Okazaki ◽

Yoshiharu Fujii

Keyword(s):

Oxidative Stress ◽

Underlying Mechanism ◽

Inhibitory Effect ◽

Succinic Semialdehyde ◽

Succinic Semialdehyde Dehydrogenase ◽

Knock Out ◽

Soybean Seedlings ◽

Semialdehyde Dehydrogenase ◽

Phytotoxic Effect ◽

And Oxidative Stress

The phytotoxic effect of the allelochemical cyanamide has been well-documented yet the underlying mechanism for this phenomenon has not been fully characterized. Cognizant of the putative inhibitory effect of cyanamide on aldehyde dehydrogenases (ALDHs), we hereby show that the capacity of mitochondrial preparations from cyanamide-treated soybean seedlings to oxidize acetaldehyde and succinic-semialdehyde was dose-dependently reduced to at most 55% and 70%, respectively. Cyanamide-treated plants exhibited oxidative stress (i.e. increased lipid peroxidation and H2O2 accumulation) that was exacerbated upon exposure to UV-A – symptoms reminiscent of ALDH and succinic-semialdehyde dehydrogenase (SSADH) knock-out Arabidopsis mutants. We suggest that the inhibition of mitochondrial ALDH and SSADH may be a contributory mechanism to the burst in oxidative stress mediated by cyanamide.

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The DM Domain Transcription Factor MAB-3 Regulates Male Hypersensitivity to Oxidative Stress in Caenorhabditis elegans

Molecular and Cellular Biology ◽

10.1128/mcb.01459-09 ◽

2010 ◽

Vol 30 (14) ◽

pp. 3453-3459 ◽

Cited By ~ 8

Author(s):

Hideki Inoue ◽

Eisuke Nishida

Keyword(s):

Oxidative Stress ◽

Transcription Factor ◽

Caenorhabditis Elegans ◽

Sex Determination ◽

Target Genes ◽

Stress Sensitivity ◽

Gata Factor ◽

X Chromosomes ◽

C Elegans ◽

Dm Domain

ABSTRACT Sex differences occur in most species and involve a variety of biological characteristics. The nematode Caenorhabditis elegans consists of two sexes, self-fertile hermaphrodites (XX) and males (XO). Males differ from hermaphrodites in morphology, behavior, and life span. Here, we find that male C. elegans worms are much more sensitive than hermaphrodites to oxidative stress and show that the DM domain transcription factor MAB-3 plays a pivotal role in determining this male hypersensitivity. The hypersensitivity to oxidative stress does not depend on the dosage of X chromosomes but is determined by the somatic sex determination pathway. Our analyses show that the male hypersensitivity is controlled by MAB-3, one of the downstream effectors of the master terminal switch TRA-1 in the sex determination pathway. Moreover, we find that MAB-3 suppresses expression of several transcriptional target genes of the ELT-2 GATA factor, which is a global regulator of transcription in the C. elegans intestine, and show that RNA interference (RNAi) against elt-2 increases sensitivity to oxidative stress. These results strongly suggest that the DM domain protein MAB-3 regulates oxidative stress sensitivity by repressing transcription of ELT-2 target genes in the intestine.

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