| Paenibacillus polymyxa is widely used in the development of biological fertilizers.It is a plant rhizosphere-promoting bacteria(PGPR),which has strong rhizosphere colonization and growth-promoting capabilities.During laboratory research,a mutant of P.polymyxa SC2,SC2-M1,was isolated,which significantly changed its co lonization and growth-promoting ability,biofilm-forming ability,spore-forming ability and antibacterial substance synthesis.Athletic ability is weakened to varying degrees,which seriously affects the colonization ability of P.polymyxa SC2 and its application in agricultural production.In the genomic and transcriptome analysis of the mutant SC2-M1,a point mutation in an important gene,spo0 A,was found.In order to study the role of spo0 A gene in colonization and growth promotion ability,we compared the protein between mutant strain SC2-M1 and complement strain SC2-M1(spo0A)by relative quantitative proteomics,and explored the metabolic pathways involved in spo0 A gene.The spo0 B gene was knocked out and over-expressed to investigate the regulatory role of the two-component system spo0B-spo0 A in the metabolic pathway.Through liquid chromatography-tandem mass spectrometry(LC-MS/MS)analysis,a total of 2852 proteins were identified.A database search and proteomic analysis of these proteins were performed.The analysis results revealed a total of 452 differentially expressed proteins.There are 306 kinds of proteins were significantly up-regulated,and 146 kinds of proteins are significant down-regulations.C luster analysis of these differential proteins found that most of the differential proteins are closely related to cell movement and metabolism,carbohydrate metabolism and transport,and they were consistent with which we have detected the difference traits,such as the loss of motility,biological model formation,spore-forming ability.In addition,71 proteins with unknown functions were found.These proteins may be involved in undiscovered metabolic pathways regulated by the spo0 A gene.In the two-component system spo0B-spo0 A,we modified the spo0 B gene and constructed the spo0 B knockout strain named SC2-Δspo0B and the over-expressing strain named SC2-PHY300PLK-spo0 B.Through the ability to form biofilms,spores,and bacteria quantitative analysis of capacity revealed that in the knockout strain SC2-Δspo0B,biofilm formation was enhanced,spore formation was weakened,and bacterial antagonism was weakened;in the over-expressing strain SC2-PHY300PLK-spo0 B,biofilm formation was decreased and spore formation was reduced Enhanced,bacterial antagonis m was enhanced.The proteomics results showed that the main reason for the change in the morphology and function of the mutant SC2-M1 was the mutation of the spo0 A gene.The spo0 A gene is a global regulatory factor that regulates the expression of multiple metabolic pathways.In the experiment,It has been proven to play important roles in spore formation,biofilm formation,motility,and antimicrobial synthesis pathways.Many differently expressed unknown functional proteins were also found in the omics data.These proteins may be involved in the metabolic pathways regulated by spo0 A.They may be metabolic pathways not found in Paenibacillus polymyxa.It can provided the basis for the study of metabolic pathway in Paenibacillus polymyxa.The results of the transformation of the spo0 B gene showed that changes in the expression of Spo0 B affected the strength of the strain in spore formation,biofilm formation and bacterial antagonism.In a two-component system,the spo0 B gene and spo0 A gene coordinated to regulate the expression of colonization-related traits.This finding is of great significance for us to transform Paenibacillus polymyxa to make it more suitable for colonization in the rhizosphere.In summary,the spo0 A gene is involved in regulating the metabolic pathways of bacterial colonization-related traits.The point mutation of the spo0 A gene in the mutant SC2-M1 is the main reason for the defect.In proteomics,a total of 452 proteins with significantly different expression levels were identified,including more than 70 proteins with unknown functions.These unknown proteins provide an important basis for exploring new regulatory pathways of the spo0 A gene.In the two-component system spo0B-spo0 A,the spo0 B gene and spo0 A gene have a synergistic effec t in regulating the colonization ability of bacteria.The results show that the study provides a theoretical basis for further research on Paenibacillus polymyxa,provides important contributions for enhancing the rhizosphere colonization ability of Paenibacillus polymyxa and its wide application in agricultural production. |
