The Molecular Mechanism Of AtVIP1 Mediates IAA And ABA Signaling Pathways To Regulate Lateral Root Development In Sorghum-preliminary Study

Lateral root is the determinant of plant root structure.In addition to providing anchorage,it is also an organ for plants to explore the water and nutrients in the soil environment.Different from primary roots,lateral roots are formed after embryo and germinated in the depth of root.IAA is an essential phytohormone for the initiation and establishment of lateral roots.ABA signaling pathway can coordinate with IAA signaling pathway to regulate lateral root development.At VIP1 is a member of the b ZIP domain of Arabidopsis,which not only participates in the infection process of Agrobacterium,but also plays a role in osmotic regulation,root fluctuation,ABA response and transcription regulation.Atvip1 gene over-expressing can promote lateral root development of sorghum,but the mechanism is not clear.Therefore,it is of great significance for the growth and yield of sorghum to explore the molecular mechanism of transcription factor At VIP1 mediated IAA and ABA signaling pathway in regulating the development of sorghum lateral roots.At the same time,it is expected to obtain new key genes that affect plant root development,and provide theoretical basis for clarifying the signal pathway of plant root development.The main results are as follows:1.In order to understand the basic biological function of At VIP1,the nucleic acid sequence and amino acid sequence were analyzed by bioinformatics.Atvip1 gene was split gene,and the promoter region contains response elements such as plant hormones and stress.At VIP1 protein has many phosphorylation sites,which mainly locates in the nucleus and mitochondrial matrix.The secondary structure of At VIP1 was mainly composed of irregular curl and α-helix.Therefore,the VIP1 protein of sorghum and Arabidopsis were closely related,and the b ZIP domain were highly conserved.2.In order to improve the genetic transformation efficiency of sorghum,through Agrobacterium mediated genetic transformation of sorghum immature embryos,a total of 14Atvip1 gene over-expressing sorghum transgenic plants were obtained.The At VIP1-7 strain was screened out to promote the development of lateral roots.3.In order to study the effects of Atvip1 gene on the growth morphology and root development of sorghum seedlings.Wild-type sorghum p898012 and At VIP1-7 transgenic sorghum line were selected as experimental materials,observing the methods of morphologicaland growth index detection,observing the occurrence of lateral roots and lateral root primordia,measurement of root activity,the root expression pattern of At VIP1,and the effect of Atvip1 gene on root development was verified by applying 1.5 m M IAA.The results showed that the Atvip1 gene over-expressing sorghum mutant developed rapidly,the root system grew more luxuriantly,the number of lateral roots and their primordia increased significantly,and and the root activity increased.At VIP1 was expressed in roots,apices and lateral roots of sorghum.The application of IAA could advance the development of sorghum roots,increase the number of lateral roots and primordia.The growth state of wild-type sorghum root was similar to that of At VIP1-7 transgenic sorghum.It is suggested that the Atvip1 gene over-expressing can promote the growth and lateral root formation of sorghum seedlings,and At VIP1 may regulate IAA signaling pathway.4.In order to find out the difference of Atvip1 gene transcription level in the process of sorghum root development,and to screen out the key genes of IAA and ABA pathway regulating the root development,the sorghum seedlings at the stage of three leaves and one heart was used as the experimental material to sequence and analyze the transcriptome.247 differentially expressed genes(DEGs)and 235 differentially annotated genes were screened.COG analysis showed that 59 DEGs were distributed in 17 gene families,mainly carbohydrate transport and metabolism.GO analysis showed that 167 DEGs were enriched in 18 biological processes,12 cell components and 12 molecular functions,and mainly cell metabolism of biological processe.47 DEGs were enriched to 29 KEGG pathways,mainly in the pathway of phenylpropionic acid biosynthesis.5.In order to study the regulatory effect of transcription factor At VIP1 on IAA and ABA signal pathway key genes at the transcription level.we used LC-MS was used to detect IAA and ABA hormone contents,combined with transcriptome pathway data and q RT-PCR to analyze the expression patterns of key genes regulating lateral root development of IAA and ABA pathway.In sorghum roots,the over-expressing Atvip1 gene can positively regulate the gene expression of Sbyucca2 and Sbnced1,which are key enzymes of IAA and ABA biosynthesis,and cause the significant increase of IAA and ABA contents.The Atvip1 gene over-expressing positively regulated the transcriptional expression of Sbmpk3,Sbmyb44 and Sbvp1 gene,and affected the transcription of Sbvip1 gene in roots.Sbiaa14 and Sbarf7 and Sblbd16 gene,which the key genesin IAA signaling pathway,were up-regulated significantly.The expression of ABA signal receptor Sbpyl8 gene and negative regulatory factor Sbpp2c5 gene was up-regulated,while the expression of positive regulatory factor Sbsnrk2 gene was down-regulated.After exogenous IAA treatment,the expression of Sbvip1,Sbvp1,Sbarf7 and Sbmyb44 genes were up-regulated,and the change trend was similar to that the Atvip1 gene over-expressing sorghum,and the expression of Sbpyl8 gene had no change.According to bioinformatics analysis,it was found that the promoter regions of myb44 and vp1 genes in sorghum not only had b ZIP targeting sequences,but also had specific response motifs of VIP1 transcription factors.It was speculated that At VIP1 could regulate the transcription of Sbmyb44 and Sbvp1 genes.It is suggested that the Atvip1 gene over-expressing may promote sorghum lateral roots development by negatively regulating ABA-Sn RK2 signaling pathway and positively regulating auxin signaling pathway at the transcription level.In conclusion,the transcription factor At VIP1 regulates the IAA and ABA signaling pathways at the transcription level to promote the growth and development of sorghum lateral roots.