Functional Analysis Of SbCrRLK1L In Sweet Sorghum(Sorghum Bicolor L.) Under Aluminum Stress

Soil acidification is a major problem that restricts agricultural development.Aluminum（Al）toxicity is the main limiting factor for crop yield in acid soils.Under acidic soil conditions（p H<5.0）,Al³⁺dissolved in the soil solution is highly toxic to plants,inhibiting the growth of plant roots,leading to underdevelopment of roots,which directly affects the ability of plants to absorb water and mineral nutrients.Plants need to sense and integrate a large number of extracellular stimuli,and influence complex regulatory pathways such as transcription level,protein level and metabolism level through signal transduction,and ultimately regulate their own growth and development.Proteins synthesized by transcription and translation require a series of post-translational modifications.Among them,the phosphorylation modification catalyzed by protein kinases plays a key role in the process of cell life activities.Receptor-like kinases（RLKs）are a subfamily of protein kinases,which play an important role in the Al tolerance process of plants.Overexpression of receptor-like kinases WAK1 and RLKx in Arabidopsis can improve the Al tolerance of Arabidopsis.In addition,Cr RLK1L receptor kinases can support the growth of plant roots under the stress of Cu,Cd,and Ni metal ions.The early stage of the laboratory has been devoted to exploring the Al toxicity mechanism and Al tolerance mechanism of sweet sorghum.However,there are few reports about the sweet sorghum receptor kinase SbCrRLK1Ls so far.In this experiment,sweet sorghum Al-tolerant variety ROMA was used as material.Through bioinformatics analysis,expression pattern analysis,subcellular localization,tissue localization analysis,and phenotypic analysis of heterologous expression in Arabidopsis,the function of sweet sorghum SbCrRLK1Ls was explored,and the specific research results were as follows.1.Using the amino acid sequences of Cr RLK1,At HERK1 and Os HERK1 to perform BLAST analysis,a total of 16 SbCrRLK1L receptor kinases were obtained.Amino acid sequence alignment and phylogenetic analysis showed that all 16SbCrRLK1L had Malectin-like domains and serine/threonine kinase domains.Among them,SbCrRLK1L1,SbCrRLK1L2 and At HERK1 had high homology;SbCrRLK1L9,SbCrRLK1L10 and At HERK2 were homologous in high degree;SbCrRLK1L11,SbCrRLK1L13 and SbCrRLK1L16 had high homology with At FER.2.Expression pattern analysis showed that the expression of 16 SbCrRLK1L genes at the transcriptional level was induced by Al.With the increasing of Al treatment time,the expression pattern of SbCrRLK1Ls could be divided into 3 types（Type I,II,III）;with the increasing of Al treatment concentration,the expression pattern of SbCrRLK1Ls can also be divided into 3 types（Type A,B,C）;It is speculated that SbCrRLK1Ls may play different functions in Al stress response.SbCrRLK1Ls were expressed in varying degrees in the roots and shoots of sweet sorghum.For most of sweet sorghum SbCrRLK1Ls,after Al treatment,the relative expression of SbCrRLK1Ls in the root tip 0-1 cm increased significantly.3.The 7 genes in the SbCrRLK1Ls family with high homology to At HERK1,At HERK2 and At FER were cloned,and SbCrRLK1L1（the homologous gene of At HERK1）,SbCrRLK1L10（the homologous gene of At HERK2）,and SbCrRLK1L16（the homologous gene of At FER）were successfully cloned.The subcellular localization results showed that the three SbCrRLK1L proteins were all localized in the plasma membrane.4.GUS staining tissue location analysis showed that the genes SbCrRLK1L1,SbCrRLK1L10 and SbCrRLK1L16 were expressed to varying degrees in the roots and shoots of the plant,regardless of Al treatment.The genes SbCrRLK1L1 and SbCrRLK1L16 had stronger GUS activity in the shoots（mainly leaves）,but weaker in roots.Al stress increased the GUS activity of these two genes in leaves.In contrast,the gene SbCrRLK1L10 has stronger GUS activity in roots and weaker activity in shoots.5.Phenotypic analysis of SbCrRLK1L1,SbCrRLK1L10 and SbCrRLK1L16heterologously expressed in Arabidopsis wild-type showed that under Al stress,overexpression of SbCrRLK1L1 and SbCrRLK1L10 could improve the Al tolerance of transgenic Arabidopsis,while SbCrRLK1L16 couldnot improve Al resistance of transgenic Arabidopsis.