The Role Of Calcium On Wheat Hypersensitive Reaction Induced By Wheat Leaf Rust Fungus

In the experiment wheat cultivar Lovrin10 and different leaf rust fungus races formed compatible and incompatible interactions. Hypersensitive reaction (HR) is an important factor to resist leaf rust infection in incompatible interaction. Firstly we carried pharmacology experiment.Calcium chelator(EGTA) and calcium channel blocker (LaCl₃ and Verapamil) were separately injected into wheat leaves before inoculation to study their effects on HR.The results showed that three reagents could obviously reduce the area of dying cells caused by HR.The higher the concentration,the smaller the area of dying cells;while calcium ionophore A23187 was injected into wheat leaves without inoculation, HR could be detected.The HR area increased with A23187 concentration,but they were not direct proportion.So we deduced that Calcium may play important role in signal transduction of wheat to resist the infection of leaf rust fungus.Using potassium pyroantimonate precipitate method and transmission electron microscope technique, subcelluar distribution of soluble calcium in wheat mesophyll cells were observed in different infection phases.At the same time/we comparatively observed ultrastructural changes of wheat mesophyll cells at different time points in different compatible combination.The results are given as following:1.In the wheat mesophyll cells without inoculation,calcium-containing pyroantimonate deposition form big granule and ruleless obturation middle empty objection in intercellular space;while calcium-containing pyroantimonate deposition are small and assemble together in vacuole.The distribution trend of calcium is the same in different time of sampling.These observation suggested that there are a lot of soluble calcium in intercellular space and vacuole, suggesting that cytosolic calcium concentration was maintained stabilizing level under normal state.Nuclei were round in shape and chromatin distributed evenly;Chloroplasts are at the edge of cells;Mitochondrion structure is normal.2.In incompatible interaction,the cytoplasm swarmed with mass calcium from intercellular space at the early stage of wheat-leaf rust interaction(after inoculation 4-12 hours),thus calcium deposition increase in cytoplasm. 12 hours after inoculation,the chloroplast became disordered,nuclei gradually became irregular;24-72 hours afterinoculation,a large amount of calcium depositions were found in host HR cells touched by infection hyphae;Mitochondria gradually became empty;96-144 hours after inoculation,calcium depositions distributed in intercellular space,HR cell membrane looks integrated.There are more darker and collapsed components in cells;At last HR cells disappeared and intercellular space became larger.3.hi compatible interaction, the rise of cytosolic calcium concentration were later than incompatible combinations.The size and quantity of calcium-containing pyroantimonate deposition granules were less than that in incompatible combination.ln the late phase of interaction^ little calcium deposition distributed on the cell membrane and the cell wall.Ultrastuctural changes were later than that in incompatible interaction and difference were remarkable.48 hours after inoculation,nuclei showed a little changes and collapsed until 144 hours;Although chloroplast became out of shape, a lot of starches in them still could be observed in 120 hours;the mitochondria shrank and became disorganized.4.1n incompatible combination,the hyphae in intercellular were very less and haustorial mother cells and haustorial structure could scarely be seen,suggesting hyphae growth is restrained;while in compatible interaction,intercellular hyphae have more branches and more haustorial mother cells,haustorial structure were seen frequently in infected cells,suggesting hyphae growth is normal.In the early stage(96 hours after inoculation),the cell structure slightly deformed,suggesting that the cells had dying trend,but this death was slow and passive,so hyphaes could get nutrition and grow normally.To sum up all above mentioned results, it is concluded that cytosolic calcium increase mainly depends calcium influx;calcium is necessary for HR.Thus these results provided more powerful proof for calcium involving in HR signal transduction in wheat-leaf rust fungus interaction.