Mdivi-1 Induced Mitochondrial Fusion as a Potential Mechanism to Enhance Stress Tolerance in Wheat

Abstract

Mitochondria play a key role in providing energy to cells. These organelles are constantly undergoing dynamic processes of fusion and fission that change in stressful conditions. The role of mitochondrial fusion in wheat root cells was studied using Mdivi-1, an inhibitor of the mitochondrial fragmentation protein Drp1. The effect of the inhibitor was studied on mitochondrial dynamics in the roots of wheat seedlings subjected to a wounding stress, simulated by excision. Treatment of the stressed roots with the inhibitor increased the size of the mitochondria, enhanced their functional activity, and elevated their membrane potentials. Mitochondrial fusion was accompanied by a decrease in ROS formation and associated cell damage. Exposure to Mdivi-1 also upregulated genes encoding the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and an energy sensor AMP-dependent protein sucrose non-fermenting-related kinase (SnRK1), suggesting that mitochondrial fusion is associated with a general activation of energy metabolism. Controlling mitochondrial fusion rates could change the physiology of wheat plants by altering the energy status of the cell and helping to mitigate the effects of stress.

Keywords: energy status; fusion; mitochondria; redox metabolism; wheat; wounding.

Figure 1

Ultrastructure of mitochondria in wheat roots: (a) orthodox mitochondria of oval shape in control roots (1 h); (b) enlarged mitochondrium in roots treated with 0.1 μM Mdivi-1 (1 h); (c) mitochondrium of irregular shape in Mdivi-1 treated roots (1 h). In the increased fragment, the arrows indicate constriction zones. cw—cell wall, er—endoplasmic reticulum, m—mitochondrium, n—nucleus, v—vacuole. Scale bar is 0.5 μm.

Figure 2

Mitochondrial membrane potential visualized with TMRM: (a,c,e)—control for 1, 3, 5 h, respectively; (b,d,f)—roots treated with 0.1 μM Mdivi-1 for 1, 3, 5 h, respectively. Scale bar is 50 µm.

Figure 3

Oxygen consumption of wheat roots treated with 0.1 μM Mdivi-1. Control—white squares, Mdivi-1—black circles. Treatment with Mdivi-1 had a significant effect, p ˂ 0.05.

Figure 4

Content of H₂O₂ (A) and malondialdehyde (MDA) (B) in wheat roots following treatment with 0.1 μM Mdivi-1.

Figure 4

Content of H₂O₂ (A) and malondialdehyde (MDA) (B) in wheat roots following treatment with 0.1 μM Mdivi-1.

Figure 5

Autophagosome formation visualized with LysoTracker Red: (a,c,e)—control for 1, 3, 5 h, respectively; (b,d,f)—roots treated with 0.1 μM Mdivi-1 for 1, 3, 5 h, respectively. Scale bar is 50 µm.

Figure 6

Gene expression of wheat roots cells treated with 0.1 μM Mdivi-1: (A)—isoforms of glyceraldehyde-3-phosphate dehydrogenase (GAPDH); (B)—subunits of sucrose non-fermenting-related kinase (SnRK1); (C)—isoforms of ATG8 genes. Grey bar—intact roots, white bars—control roots for 1, 3, 5 h, black bars—roots treated with 0.1 μM Mdivi-1 for 1, 3, 5 h. * p ≤ 0.05; ** p ≤ 0.005.