Effect Of PfPMI2 Gene On The Changes Of Gene Expression In Populus Trichocarpa

PMI2,a member of the WPR gene family,is a key gene which can regulate chloroplast movement and plays an important role in regulating leaf yellowing.Paulownia witches’ broom(Pa WB)is an infectious disease caused by phytoplasma infection,which can cause yellowing of leaves.Our previous study found that the expression level of PfPMI2 was significantly up-regulated during the occurrence of Pa WB,suggesting that this gene may play an important role in the yellowing leaves during the occurrence of PaWB.Therefore,the effect of PfPMI2 on the morphology and gene expression of Populus trichocarpa were explored by using bioinformatics,genetic transformation,transcriptome,and small RNA sequencing.The main results are summarized as follows:1.The PfPMI2 gene is relatively conserved in evolution and has tissue expression specificity.PfPMI2 has a conserved and stably inherited amino acid sequence during evolution,and may have highly similar functions in different species.In the amino acid composition of PfPMI2 protein,the content of glutamic acid is the largest,followed by lysine and leucine,and the content of cysteine is the least,only one.PfPMI2 was significantly expressed in the roots,stem segments and leaves of Paulownia alba,but the expression level in leaves was higher than that in stem segments and roots.2.Obtained PfPMI2 transgenic plants of P.trichocarpa.The pART-CAM-FLAG-PfPMI2 overexpression vector was constructed,and transgenic plants were obtained.Morphologically,the PfPMI2 transgenic plants showed yellowed leaves.Compared with wild-type plants,the content of chlorophyll a in PfPMI2 transgenic plants was reduced,the content of chlorophyll b was basically unchanged,and the content of superoxide anion was significantly increased.3.Screened out the genes that respond to the expression of PfPMI2.In the transcriptome sequencing of PfPMI2 transgenic plants,an average of 6.75 G of raw data was obtained.The average alignment rate of the sequencing data to the genome was 94.07%,and the average alignment rate to the gene set was 83.39%.A total of29096 genes were detected.Compared with the wild-type plants,2981 were differentially expressed in PfPMI2 transgenic plants.GO enrichment analysis showed that the differential genes were distributed in 91 GO items;KEGG enrichment analysis showed that the differential genes were annotated into 117 KEGG pathways;the differential genes contained a total of 142 transcription factors,which belonged to25 gene families.4.The miRNAs expressed in response to PfPMI2 were screened and their target genes were identified.An average of 32.42 M data was obtained in two libiaries,and the average alignment rate of the sequencing data to the genome was 90.56%,and a total of 358 small RNAs were detected.Compared with the wild-type plants,differentially expressed miRNAs were screened in PfPMI2 transgenic plants,which targeted 245 m RNAs,Of which,142 target genes were annotated to GO cellular components,155 genes were annotated to molecular functions,and 107 genes were annotated to biology process;KEGG enrichment analysis found that 35 of the target genes were annotated to 31 KEGG pathways.5.MiRNA-mRNA regulatory modules were discovered in response to PfPMI2 expression.Correlation t analysis of ranscriptome and miRNA revealed 25 target genes of the differentially expressed miRNA response to the expression of PfPMI2,and these genes were mainly involved in the regulation of fatty acid elongation,brassinolide biosynthesis,terpenoid backbone biosynthesis,ubiquitin-mediated induced proteolysis,phenylpropane biosynthesis,plant-pathogen interactions,cell cycle and antibiotic biosynthesis pathways.Together,PfPMI2 may cause leaf curling and yellowing symptoms in P.trichocarpa transgenic plants by affecting the related miRNA-m RNA regulatory modules in the above metabolic pathways.