The Function Of JMJ21 In The Regulation Of Shoot Apical Meristem Development In Arabidopsis

Shoot apical meristem(SAM)are responsible for elaborating the above ground portions of the plant,maintenance of SAM activity depends on the coordination between the rates of cell differentiation and proliferation,understanding the machanism of SAM development plays a key role in plant development.We identified a T-DNA insertion mutant jmj21,showing distinguishable smaller SAM than wild type.Histone demethylase activity analysis demonstrated that JMJ21 might lack the enzymatic activity,which is different from other JMJ family members.RNA-Seq data revealed down-regulation of WUS and STM in jmj21 mutant as compared with wild type,which are associated with defective SAM development.The molecular mechanism of JMJ21-mediated SAM development was preliminary uncovered in this study.The main results are as follows:(1)A T-DNA insertion mutation of JMJ21 caused a pleiotropic phenotype compared with wild type,including smaller seedlings,dwarf plants and fewer branches.The jmj21 mutant displayed significant smaller SAM than the wild type.JMJ21 was highly expressed in SAM,and JMJ21 protein was localized at nucleus.(2)JMJ21 belongs to Jumonji C(Jmj C)Domain-containing(JMJ)protein family,a large family of JMJ proteins has been discovered as histone demethylases.It can influence gene expression via histone demethylation process.Histone demethylase activity of JMJ21 was verified to uncover the mechanism of JMJ21 in regulating gene expression.Surprisingly,JMJ21 was found lacking enzymatic activity at H3R2,H4R3,H3K4,H3K9,H3K27 and H3K36 level.Amino acid sequence alignment between JMJ21 and other two homologous proteins(Arabidopsis JMJ22 and human JMJD6),which have been previously reported to have histone demethylase activity,showed that the first conserved cofactor ?-ketoglutarate(?-KG)binding residue Alanine of JMJ21 was different from Serine in Arabidopsis JMJ22 and Threonine in human JMJD6.(3)RNA-Seq was performed to investigate the mechanism of JMJ21 in regulating SAM development.Data analysis indicated that JMJ21 regulated SAM development through repressing expression of three critical genes,WUSCHEL(WUS),SHOOTMERISTEMLESS(STM)and CLAVATA 3(CLV3).Genetic analysis between JMJ21 and WUS,STM showed that jmj21 wus-am and jmj21 stm-2 double mutants lacked a primary shoot meristem,consistent with wus-am and stm-2 single mutant.These results indicated that JMJ21 acted upstream of WUS and STM to regulate SAM development.(4)SHEPHERD(SHD)was identified from differentially expressed genes in jmj21 mutant from RNA-Seq data.SHD was reported to play a key role in SAM development,which was localized in the endoplasmic reticulum(ER)and was important for the correct folding of CLV3 protein.shd mutant showed expanded SAM,functional CLV3 protein accumulation was disturbed,leading to the increase of WUS and CLV3.q RT-PCR and in situ hybridization results showed that SHD expression level was increased in jmj21 mutant.Thus we considered that JMJ21 regulated SAM development through affecting the expression of SHD.In this study,we uncovered the molecular mechanism of JMJ21 in regulating the development of shoot apical meristem,which provided new data for the further improvement of SAM development regulation network.