Optimization of the HyPer sensor for robust real‐time detection of hydrogen peroxide in the rice blast fungus

Huang, Kun; Caplan, Jeff; Czymmek, Kirk J.; Donofrio, Nicole

doi:10.1111/mpp.12392

Cited by 14 publications

(10 citation statements)

References 38 publications

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“…The use of HyPer has been concentrated in measuring levels of H 2 O 2 in living cells under receptor activation and fisiopathological conditions both in mammalian [12] , [13] , [14] and plant cells [15] , [16] , [17] . Intracellular H 2 O 2 levels are driven primarily from enzymatic reductions of superoxide anions, carried out by superoxide dismutases (SODs).…”

Section: Introductionmentioning

confidence: 99%

Insights into the HyPer biosensor as molecular tool for monitoring cellular antioxidant capacity

et al. 2018

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Aerobic metabolism brings inexorably the production of reactive oxygen species (ROS), which are counterbalanced by intrinsic antioxidant defenses avoiding deleterious intracellular effects. Redox balance is the resultant of metabolic functioning under environmental inputs (i.e. diet, pollution) and the activity of intrinsic antioxidant machinery. Monitoring of intracellular hydrogen peroxide has been successfully achieved by redox biosensor advent; however, to track the intrinsic disulfide bond reduction capacity represents a fundamental piece to understand better how redox homeostasis is maintained in living cells.In the present work, we compared the informative value of steady-state measurements and the kinetics of HyPer, a H2O2-sensitive fluorescent biosensor, targeted at the cytosol, mitochondrion and endoplasmic reticulum. From this set of data, biosensor signal recovery from an oxidized state raised as a suitable parameter to discriminate reducing capacity of a close environment. Biosensor recovery was pH-independent, condition demonstrated by experiments on pH-clamped cells, and sensitive to pharmacological perturbations of enzymatic disulfide reduction. Also, ten human cell lines were characterized according their H2O2-pulse responses, including their capacity to reduce disulfide bonds evaluated in terms of their migratory capacity.Finally, cellular migration experiments were conducted to study whether migratory efficiency was associated with the disulfide reduction activity. The migration efficiency of each cell type correlates with the rate of signal recovery measured from the oxidized biosensor. In addition, HyPer-expressing cells treated with N-acetyl-cysteine had accelerated recovery rates and major migratory capacities, both reversible effects upon treatment removal. Our data demonstrate that the HyPer signal recovery offers a novel methodological tool to track the cellular impact of redox active biomolecules.

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Section: Introductionmentioning

confidence: 99%

Insights into the HyPer biosensor as molecular tool for monitoring cellular antioxidant capacity

et al. 2018

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“…Subsequently, a more sensitive HyPer2 sensor was developed to measure redox balance in mutants impaired for oxidative stress regulation in F. graminearum (Mentges and Bormann, 2015). This sensor has been further codon optimized from Neurospora crassa for use in M. oryzae and other fungi (Huang et al, 2016). The MoHyPer sensor tracks ROS during the development of M. oyzae on artificial hydrophobic surfaces and in planta.…”

Section: Hyper Redox Sensorsmentioning

confidence: 99%

“…A genetically encoded HyPer (hydrogen peroxide) sensor used in filamentous fungi-originally developed to detect intracellular ROS in mammalian cells using a circularly permuted yellow fluorescent protein (cpYFP) bound to the regulatory domain of the H 2 O 2 -sensing OxyR protein in Escherichia coli (Belousov et al, 2006)-has been optimized for several fungal phytopathogens (Huang et al, 2016;Ronen et al, 2013). The HyPer sensor allows for intracellular measurement of ROS and is specific and sensitive to H 2 O 2 (Belousov et al, 2006).…”

Section: Hyper Redox Sensorsmentioning

confidence: 99%

Reactive oxygen species metabolism and plant-fungal interactions

Segal

Wilson

2018

Fungal Genetics and Biology

155

103

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Fungal interactions with plants can involve specific morphogenetic developments to access host cells, the suppression of plant defenses, and the establishment of a feeding lifestyle that nourishes the colonizer oftenbut not always-at the expense of the host. Reactive oxygen species (ROS) metabolism is central to the infection process, and the stage-specific production and/or neutralization of ROS is critical to the success of the colonization process. ROS metabolism during infection is dynamicsometimes seemingly contradictory-and involves endogenous and exogenous sources. Yet, intriguingly, molecular decision-making involved in the spatio-temporal control of ROS metabolism is largely unknown. When also considering that ROS demands are similar between pathogenic and beneficial fungal-plant interactions despite the different outcomes, the intention of our review is to synthesize what is known about ROS metabolism and highlight knowledge gaps that could be hindering the discovery of novel means to mediate beneficial plant-microbe interactions at the expense of harmful plant-microbe interactions.

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“…However, it is difficult to use these methods to track ROS generation and accumulation during rice– M. oryzae interaction. It might be helpful to use reporters, such as the luciferase reporter, roGFP, or the Hyper sensor, for living cell imaging to answer this important question [ 19 , 47 , 109 ]. Another important question is why the ROS burst actually occurs.…”

Section: Discussionmentioning

confidence: 99%

Every Coin Has Two Sides: Reactive Oxygen Species during Rice–Magnaporthe oryzae Interaction

Kou

Qiu

Zeng

2019

IJMS

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Reactive oxygen species (ROS) are involved in many important processes, including the growth, development, and responses to the environments, in rice (Oryza sativa) and Magnaporthe oryzae. Although ROS are known to be critical components in rice–M. oryzae interactions, their regulations and pathways have not yet been completely revealed. Recent studies have provided fascinating insights into the intricate physiological redox balance in rice–M. oryzae interactions. In M. oryzae, ROS accumulation is required for the appressorium formation and penetration. However, once inside the rice cells, M. oryzae must scavenge the host-derived ROS to spread invasive hyphae. On the other side, ROS play key roles in rice against M. oryzae. It has been known that, upon perception of M. oryzae, rice plants modulate their activities of ROS generating and scavenging enzymes, mainly on NADPH oxidase OsRbohB, by different signaling pathways to accumulate ROS against rice blast. By contrast, the M. oryzae virulent strains are capable of suppressing ROS accumulation and attenuating rice blast resistance by the secretion of effectors, such as AvrPii and AvrPiz-t. These results suggest that ROS generation and scavenging of ROS are tightly controlled by different pathways in both M. oryzae and rice during rice blast. In this review, the most recent advances in the understanding of the regulatory mechanisms of ROS accumulation and signaling during rice–M. oryzae interaction are summarized.

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Optimization of the HyPer sensor for robust real‐time detection of hydrogen peroxide in the rice blast fungus

Cited by 14 publications

References 38 publications

Insights into the HyPer biosensor as molecular tool for monitoring cellular antioxidant capacity

Insights into the HyPer biosensor as molecular tool for monitoring cellular antioxidant capacity

Reactive oxygen species metabolism and plant-fungal interactions

Every Coin Has Two Sides: Reactive Oxygen Species during Rice–Magnaporthe oryzae Interaction

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