Cell death in plant protection: improving the security of food supply by the means of molecular biology

DOI

Received: October 2022 – Accepted: November 2022

¹ Budapest University of Technology and Economics, Laboratory of Biochemistry and Molecular Biology, Department of Applied Biotechnology and Food Science

Keywords

programmed cell death, harpin proteins, acrolein, ferroptosis-like cell death, heat stress

2. Introduction

Protecting plants from biotic stress caused by pathogens and abiotic stress caused by adverse environmental effects is an important and increasing challenge for agriculture. Therefore, it has become important to understand the background biochemistry better. Thus, the development of new, more effective ways of plant protection and the enhancement of their stress tolerance could also be possible, thereby increasing the security of food supply.

3. Harpins in plant protection

Chemical pesticides can cause long-term and severe environmental effects. Therefore, efforts are being made to replace them with environmentally friendly biopesticides. One promising avenue for biopesticide development could be the use of the elicitors named harpin proteins. Harpins are acidic, heat-stable, cysteine-poor but glycine- and leucine-rich proteins produced by plant-pathogenic bacteria with a type III secretion system, such as Pseudomonas syringae and Erwinia amylovora [1, 2, 3]. When purified harpins are applied directly to plants at low concentrations (~100 nM) or expressed in plant cells, they induce the expression of immune response genes that make the plant more resistant to pathogens. In addition, other beneficial effects have been described in harpin-treated plants, such as higher yields and more intensive growth [4].

The exact cause of the beneficial effects of harpins is currently elusive. However, earlier results from our research group suggest that harpin treatment can induce biotic stress in Arabidopsis thaliana suspension cell cultures. This results in a so-called oxidative burst. In the course of this phenomenon the amount of reactive oxygen species (ROS) suddenly and significantly increases, which induces the antioxidant system of the cells. Our data indicated that the expression of VTC2 and 5 enzymes, which catalyze the rate-determining step of ascorbic acid biosynthesis, were increased. The expression and activity of L-galactono-1,4-lactone dehydrogenase (GLDH), that catalyzes the final step of the biosynthesis, were also increased. Presumably, as a result of the above, the ascorbic acid content of the harpin-treated cells was higher than that of non-treated cells. The activity of the enzymes of the ascorbate-glutathione cycle, which is a central element of the plant antioxidant system, is also enhanced [5]. Based on these, harpins can induce biotic stress in plants without an actual pathogen infection, which, among other things, induces the plant antioxidant system, thereby enhancing resistance to real pathogens.

4. The ferroptosis-like cell death

Harpin treatments at higher concentrations (>250 nM) induce hypersensitive response (HR) in plants [3, 4, 5]. HR in nature occurs when a plant is resistant to a particular pathogen (incompatible interactions). In this case, the immune system of the plant recognizes the pathogen and destroys its own attacked cells in a quick, programmed manner to protect the plant as a whole. In 2019, Dangol et al. reported that the HR during the incompatible reaction between rice (Oryza sativa) and the fungus Magnaporthe oryzae is characterized by lipid peroxidation and accumulation of ROS and ferric ions. However, treatment of the cells with the iron chelator deferoxamine or the lipophilic antioxidant ferrostatin-1 prevented the iron-dependent ROS accumulation and lipid peroxidation, leading to the complete attenuation of the HR cell death. On the base of these observations, Dangol et al. hypothesize that the so-called ferroptosis-like cell death is involved in incompatible plant-pathogen interactions [6].

Ferroptosis is a form of iron-dependent, caspase-independent programmed cell death in mammalian cells that was first described in 2012 [7]. Ferroptosis has unique morphological and biochemical features that differ from other forms of cell death, and its specific inducers (e.g. erastin and RSL3) and inhibitors (e.g. ferrostatin-1) have also been identified. The cell death process can be triggered by the depletion of cellular glutathione by erastin, or by the inhibition of the enzyme glutathione peroxidase 4 (GPX4), which plays a key role in the elimination of lipid peroxides. Regardless of the mode of induction, the initiated process is characterized by increased ROS production, lipid peroxidation and elevated cellular labile iron pool. Due to these properties, ferroptosis can be inhibited by lipophilic antioxidants (e.g. ferrostatin-1, liproxstatin-1, α-tocopherol) and iron chelators (e.g. deferoxamine). [8].

In 2017, Distéfano et al. described a cell death process in plants that is very similar to ferroptosis [9]. They treated Arabidopsis thaliana root hairs at 55 °C for 10 minutes and found that heat stress induced a decrease in glutathione levels, the shrinkage of mitochondria – a unique morphological marker of ferroptosis in mammalian cells – and cell death. The authors also showed that the rate of heat stress-induced cell death was significantly reduced in root hairs pretreated with the ferroptosis inhibitors ferrostatin-1 or ciclopirox olamine (iron chelator). However, the same inhibitors could not prevent cell death caused by 77 °C, H2O2 treatment or salt stress. The inhibitors also did not affect reproductive or developmental cell death. Based on these findings, the authors conclude that cell death induced by moderate heat stress is a unique process in plants and have named it ferroptosis-like cell death based on its high similarity to ferroptosis.

However, it is important to point out that ferroptosis-like cell death, in contrast to the caspase-independent ferroptosis of mammalian cells, appears to be a caspase-like protease-dependent process (Figure 1) [9, 10, 11].

5. Outlook

As the average temperature of the Earth rises, heat stress and heat stress-induced ferroptosis-like cell death are expected to play an increasingly important role in the life and death of plants. By studying and better understanding this cell death process, we may be able to develop targeted defences against it and thus increase the heat stress tolerance of our crops.

One may wonder how thermotolerant desert plants protect themselves against ferroptosis-like cell death. For example, the leaves of the creosote bush (Larrea tridentata), which grows in the deserts of the USA and Mexico, contain high levels of nordihydroguaiaretic acid (NDGA), a pan-lipoxygenase inhibitor lignan. It has been described earlier that lipoxygenases play an important role in ferroptosis through the enzymatic catalysis of lipid peroxidation. Thus, lipoxygenase inhibitors are expected to have an inhibitory effect on the cell death process. The inhibitory effect of NDGA on ferroptosis has already been demonstrated in mammalian cells, but has not yet been tested in plants [13,14].

As mentioned above, ferroptosis-like cell death is also involved in the HR during some pathogen attacks. Thus, a better understanding of ferroptosis-like cell death may be highly important for developing more effective biopesticides, hereby our research may help in the defence against both abiotic and biotic stresses.

6. Acknowledgement

This research was funded by the National Research, Development, and Innovation Fund of Hungary under Grants K 123752, 2018-1.2.1-NKP-2018-00005, RRF-2.3.1-21-2022-00015, and TKP2021-EGA-02.

7. References

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