Study On The Calcium Signal During The Interaction Of TcLr19 And Leaf Rust Fungus

According to our prior results on other wheat species,hypersensitive reaction(HR) played an important role in resistance response between incompatible interactions.The incompatible combination including wheat near isogenic line TcLr19 and leaf rust race 366 was studied in this experiment to reveal the mechanism further.Pharmacology experiments were carried out first:different concentrations of calcium chelator(EGTA) and calcium channel blocker on plasma membrane(Verapamil and LaCl₃) were injected into wheat leaves separately 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,and the higher the concentration,the smaller the area of dying cells.In addition to that,while calcium ionophore A23187 was injected into wheat leaves without inoculation,HR could be detected and the changing tendency of HR area increased with A23187 concentration,but there was not a linear relationship was found between them. These results suggested that calcium may play important role in signal transduction of wheat resistance to leaf rust fungus.In order to determine the change of calcium distribution in wheat mesophyll cells at early time points of infection,potassium pyroantimonate precipitate method and transmission electron microscope technique(TEM) were used during next experiment to observe the subcelluar localization of soluble calcium directly.And we also observed comparatively ultrastructural changes of wheat mesophyll cells at the same time points that we mentioned before.In the wheat mesophyll cells without inoculation,calcium-containing pyroantimonate deposition was flocculent and abundant in vacuole.It suggested that calcium concentration was at a higher level in vacuole,but slight calcium-containing pyroantimonate deposition were observed in intercellular space,chloroplast,cell wall and nuclei.This result may suggest that cytosolic calcium concentration was maintained stable state.So did the organelles at the same time points.For example,from the results of TEM,nuclei were approximate round and chromatin was well distributed in it;chloroplasts were at the edge of cells,on which basal corpuscles were in order,mitochondrion structure was normal.At the early stage of TcLr19 was inoculated with leaf rust fungus,appearance and location of calcium-containing pyroantimonate deposition changed significantly:from flocculent coagulum to granular then changed into flocculent coagulum again in vacuole. And the change of cytoplasmic calcium tendency was as followed:lesser calcium-containing pyroantimonate deposition granule moved to plasma membrane and became abundant;with the inoculate time passed,more calcium-containing pyroantimonate deposition moved to the plasma membrane or around chloroplast and mitochondrion;at last,the calcium-containing pyroantimonate deposition became to granular again.This result may be concluded into a tendency that the intracellular calcium concentration increased first then decreased when wheat infected by fungus,which reflected the change of calcium at the early stage and sensitivity as second massager. Meanwhile,we observed that chloroplasts,mitochondrion,nuclei were changed;cell wall and cytoplasm were modified.These results implied us that host cells have already apperceive the pathogenic infection,and some defensive reactions were expression.Our results were similar with those prior experiments on Lovrin 10,but,in our research, calcium-containing pyroantimonate deposition's varies trend and the change of organelle were earlier than Lovrin 10.Furthermore,calcium-containing pyroantimonate deposition's position and form were different with the results of Loverin 10.In conclusion of all results mentioned above,we deduced that TcLr19's high resistance to leaf rust fungus was correlative with the initial time of defensive reaction,in other words,the calcium signal's effective was necessary.On the other hand,the change of intracellular calcium-containing pyroantimonate deposition might suggest that intracellular calcium store took part in the calcium signal's generation.