The Molecular Mechanism Of Falciphora Oryzae Resist Biotic And Abiotic Stress

Endophytes are a group of fungi that colonize healthy plant tissues for at least part of their life cycle without obvious disease in their hosts.Endophytes have many significant biological functions in the process of interaction with plants.Firstly,the colonization of endophytes can induce systemic disease resistance in plants to resist the invasion of pathogenic fungi,viruses and bacteria.At the same time,it produces toxins that protect the host from herbivores and insects.When plants are faced with abiotic stresses,such as drought or flood,heavy metals or high and low temperature,endophytes can regulate the physiological and biochemical reactions of plants to help them survive in the adverse environment.In addition,it can produce some metabolites,used in the exploitment of cancer drugs,such as taxol developed by Trichoderma taxi.Endophytes can also promote the growth of plants,yield and efficiency.This research focusing on the aspects of anti-biotic stress and anti-abiotic stress.We obtained endophytes strains through the isolation and identification from the root of wild rice,among which one dark spetate endophyte Falciphora oryzae was identified.In the previous research work,the co-culture system of Falciphora oryzae and rice has been successfully established.The mechanism of response of Falciphora oryzae to biotic and abiotic stress was studied by co-culture system.In the anti-biological stress,we mainly focus on the rice blast resistance of F.oryzae-rice symbionts,that is,how do endophytes cause systemic immune resistance after symbiosis with plants,so as to resist the infection of pathogen.Secreted proteins play an important role in the regulation of microbe-plant interactions,and CFEM protein is the basis for the establishment of the interaction.However,the research is still in the exploratory stage.This study revealed the biological functions of CFEM protein in F.oryzae-rice symbiotic system and the induced systemic resistance of rice against foliar infection of M.oryzae,clarified the localization of CFEM protein,and revealed the molecular mechanism of CFEM protein to regulate the interaction between plants and fungi.Abiotic stress is mainly concentrated in the tolerance of cadmium stress in F.oryzae-rice symbionts.At present,cadmium pollution is a very serious environmental problem,and cadmium is also common in rice grain.Therefore,we wanted to explore whether endophytes can help rice resist heavy metal stress after symbiosis with rice.This study revealed the molecular mechanism of tolerance to cadmium stresses,including the biological function of F.oryzae in promoting rice tolerance to cadmium stress,and the response of cadmium stress.By proteomics and molecular biology techniques,the key gene involved in regulating the tolerance to cadmium stress of F.oryzae was identified,the molecular mechanism of promoting the tolerance of cadmium stress in rice was further clarified.The main results are as follows:1.The effctor FoCcw14 was screened.We selected 41 secreted protein containing Lys M,Chitin,Cutinase,CFEM domain respectively after bioinformatics analysis.Then,FoCcw14 was identified as the key protein for subsequent biological function analysis according to the transient expression in N.benthamiana leaves and measurement of expression level in the early stage.2.The biological function of FoCcw14.The deletion of FoCCW14 lead to the sensitivity of the mutant strain to hyperosmotic stress.Meanwhile,FM4-64and CMAC staining showed that the number and size of vacuoles in the mutant were seriously affected.In mutants,the vacuoles seemed to be smaller and more numerous which clustered or distributed randomly in the hyphae,and it is difficult to form larger vacuoles.3.The colonization pattern of FoCcw14 in rice roots.?FoCcw14 showed the decrease in colonization of rice roots.After 15 d colonization,we found that a large number of hyphae of wild-type were concentrated on the root hair area,especially at the base of the root hair,and then entered the epidermis and cortex of rice.Whereas,in?FoCcw14-rice symbiosis system,the number of hyphae in the root hair area were significantly reduced Transversely,in?FoCcw14-rice symbiosis system,most of the hyphae were concentrated in the epidermis and little invaded the cortex.4.FoCcw14 affects the systemic resistance of rice.The symbiosis systems were sprayed with the conidial suspension of rice blast.The leaves of ?FoCcw14-rice symbiosis system appeared typical spindle-like lesions. While the necrotic flecks were small and few in F.oryzae-rice symbiosis system.The disease area rate in?FoCcw14-rice symbiosis system was 15% higher than that in F.oryzae-rice symbiosis system.The content of protective enzymes(SOD,CAT)was also reduced in the?FoCcw14-rice symbiosis system.5.The role of Os GLP2 in rice.FoCcw14 affects the systemic resistance of rice. BiFC analysis and yeast two-hybrid system were performed to identify the protein Os GLP2 that interacted with FoCcw14.The protein Os GLP2 we have found was highly expressed in the early stage of infection.In ?FoCcw14-rice symbiosis system,the Os GLP2-RNAi plants were more susceptible than the wild type.6.Absorption and accumulation of Cd by Falciphora oryzae.F.oryzae could grow under high concentration of cadmium stress.Vacuoles and cytoplasm acted as primary sites for Cd²⁺binding,sequestration and immobilization. Elevated antioxidant levels as a detoxification strategy to neutralize Cd toxicity.7.Falciphora oryzae can improve the tolerance of rice to Cd.When F.oryzae was symbiotic with rice,a large proportion of Cd might be absorbed and immobilized by the endophytic fungus,leaving a small amount of Cd²⁺that might move into roots and be translocated to shoots.8.The role of FoSyn1 in cadmium tolerance.One SNARE Syntaxin1 protein FoSyn1(g5337)was found significantly by label-free proteomics to be significantly upregulated.The deletion of FoSYN1 caused little change in the vegetative growth under cadmium stress.FoSyn1 affects chlamydospore formation,vacuole morphology and Cd accumulation.?FoSyn1-inoculated plants were stunted,exhibiting a significant decrease in shoot height,affecting the tolerance of symbionts to Cd.