Autophagy Mechanism Of Pakchoi Under Cadmium Stress

Cabbage?Brassica campestris L.ssp.Chinensis Makino?has become one of the indispensable vegetable varieties in the market.Taking the heavy metal cadmium?Cd?,which is of high concern in today's society,as an example,the study of Cd defense function by autophagy in animals is relatively clear,but little is known in plants.In this paper,Chinese cabbage was cultured for 32 days with heavy metal cadmium nutrient solution.The cadmium concentrations were 2mg·L^-1?4mg·L^-1?6mg·L^-1?8mg·L^-11 and the control group?CK?.as follows:?1?The growth,chlorophyll fluorescence,reactive oxygen species?ROS?,antioxidant enzyme activities,mesophyll cells and root cell morphology of pakchoi were studied by nutrient liquid culture.The results showed that with the increase of Cd²⁺concentration,the cadmium content in the roots of Chinese cabbage increased significantly;the roots became shorter,Leaf chlorosis,and the fresh dry weight of Chinese cabbage decreased significantly.Under cadmium stress,the growth of Chinese cabbage was inhibited and was found of autophagy;When the concentration of Cd²⁺was less than 8mg/L,the plasmolysis,mitochondrial swelling and endoplasmic reticulum structure were damaged in the root cells.In the treatment of 2,4 and 6 mg/L,the autophagy were found in the vacuoles of the Chinese cabbage roots and the number of autophagosomes increases.However,in the 8mg·L^-11 treatment group,the cell wall was severely damaged,the organelles were broken and the nucleus was disintegrated,and the number of autophagosomes was significantly reduced.This may be caused by the high concentration and long time,exceeding the cadmium tolerance range of the Chinese cabbage roots.In addition,compared with CK,the structure of mesophyll cells in the cadmium treatment group showed impaired symptoms,that is,the chloroplast structure was vacuolated,the thylakoids were reduced and the structure was thinned,the basal particles were thinned,and the cell walls were irregularly folded.At 8mg·L^-1,the cell wall disappeared and a large vacuole appeared.The chlorophyll fluorescence parameters Fm,Fv/Fm and Qp were significantly decreased,while Qp and Y?NPQ?were significantly increased,and the SPAD value was“first rise and then fall”.Both H₂O₂ and O₂^-·are on the rise,O₂^-·is larger than H₂O₂;SOD,CAT,and APX are“first rise and then fall”,and GSH-Px is significantly increased;The increase of O2-·content is caused by endogenous oxidative stress and destruction of organelles,which activates autophagic signals and produces autophagosome formation,thus indicating that cadmium response can activate autophagy in pakchoi;In the process of cadmium stress tolerance of pakchoi,exceeding a certain threshold or insufficient autophagy can cause DNA damage and programmed cell death of pakchoi.?2?The expression levels of H3K9ac,H4K5ac,H3ac,H3K4me2 and H3K9me2proteins in autophagy group and control group were detected by Western Blotting.The results showed that with the increase of cadmium concentration,the expression level of H3K9ac continued to increase in the autophagy experimental group,while the expression levels of H3K4me2,H3K9me2,H4K5ac and H3ac increased first and then decreased.?4?RNA was extracted from the Chinese cabbage roots of the autophagy group and the control group for transcriptome sequencing.The results were as follows:114.96 Gb of high quality Data was obtained from the transcriptome,and the percentage of Q30 bases reached 88.82%or more.The Clean Reads of each group were compared with the rapeseed Z11-NCBI reference genome,and the efficiency of each group was between 79.21%-82.78%.Each group of Clean Reads was compared with GO,COG and KEGG,respectively.In the GO database,there are mainly cell processes,cells and binding;in the COG database,there are mainly general function prediction only,carbohydrate transport and metabolism,and defense mechanisms;in KEGG,there are mainly plant hormone signaling transduction,Phenylpropanoid biosynthesis,plant pathogen interaction,etc.The results of comparing the selected autophagy differential genes with the three major databases are as follows:the results in the GO database are consistent with the above;in the COG data annotation is mainly related to signal transduction mechanism,general functional prediction only and carbohydrate transport and metabolism;in the KEGG database annotation is mainly related to endocytosis and regulation of autophagy.Comparison of differential gene annotation and autophagy differential gene annotation revealed that:1.The GO database annotation results indicated that autophagy genes participate in general metabolic processes.2,autophagy pathway activation is mainly related to plant hormone signaling transduction,general function prediction only,carbohydrate transport and metabolism are closely related.