Mechanism Of XPO1-XIW1 Interaction To Regulate ABA Signaling

Abscisic acid(ABA)is an important plant stress-responsive hormone,which plays an important regulatory role in seed germination and seedling development,and can regulate the adaptability of plants to stress by initiating the expression of downstream genes.Nuclear transport of proteins is one of the important ways to regulate gene expression in cells,and is involved in processes such as plant development,abiotic stress,biological rhythm and hormone response.This study focused on the role of protein nuclear transport in regulating ABA signaling pathways and its mechanisms.Nuclear export of protein is mediated by the nuclear transporter receptor XPO1(Exportin 1).We previously identified a protein XIW1(XPO1A-Interacting WD40 protein 1)that interacts with XPO1A(Exportin 1A)by immunoprecipitation and mass-spectrometry analysis in Arabidopsis thaliana.We have proved that XIW1 expression is induced by abiotic stresses and ABA,and XIW1 interacts with ABI5(ABA Insensitive 5)in the nucleus.In this study,the XPO1-XIW1-ABI5 signal cascade was further elucidated by physiological,biochemical and molecular biological experiments.In addition,we identified and analyzed XPO1 protein in rice.The main findings are as follows:(1)The nuclear export of XIW1 was mediated by both XPO1 A and XPO1 B.Arabidopsis XIW1 localized both in the nucleus and the cytoplasm,and XIW1 interacted with nuclear transport receptors XPO1 A and XPO1 B.Mutagenesis of XPO1 A did not allow XIW1 to remain in the nucleus,suggesting that XPO1 B complemented XPO1 A function in the xpo1 a mutant,and XPO1 A and XPO1 B were functionally redundant.The XPO1 inhibitor LMB could promote XIW1 accumulation in the nucleus;(2)Phosphorylation of XIW1 is incorporated into nuclear accumulation induced by ABA.Under normal physiological conditions,XIW1 was mainly distributed in the cytoplasm.Exogenous ABA could promote the phosphorylation of XIW1 protein Thr45,Ser172 and Ser183,and promote the accumulation of XIW1 in the nucleus.Simultaneous mutation of these ABA-induced phosphorylation sites identified in this study revealed that the nuclear distribution of XIW1 was altered,indicating that ABA-induced nuclear accumulation of XIW1 was associated with phosphorylation of XIW1;(3)XIW1 affected ABI5 stability and positively regulated ABA response.In the presence of ABA,XIW1 interacted directly with ABI5 in the nucleus,which could protect ABI5 from proteasome degradation,thereby increasing ABI5 protein level and promoting ABA response.The high expression of XIW1 in the nucleus could promote the ABA response and enhanced the drought resistance of the plants;(4)Rice Os XPO1 was a developmental essential gene.Os03g0858100 was a homologous gene of Arabidopsis XPO1 in Oryza sativa.Knockout Os XPO1 gene was a lethal phenotype,which was similar to Arabidopsis XPO1 gene,xpo1 a xpo1b doublet was a lethal phenotype in Arabidopsis thaliana.Os XPO1-RNAi transgenic T0 plants were also dysplasia,and was similar to the reported Os Ran2 phenotype of transgenic rice,and both T1 seeds could not be obtained.It indicated that the interaction between Os XPO1 and Os Ran has important biological significance for the normal growth and development of plants,and also provided evidence for Os XPO1 functioned as a nuclear transporter of rice.In summary,we elucidated the XPO1-XIW1-ABI5 signaling cascade and its biological function in Arabidopsis thaliana in this study,which helps to reveal not only the biological functions of XPO1,but also the mechanism of protein nucleocytoplasmic transport in regulating ABA response.In adddition,we identified rice Os XPO1 gene,and revealed the important role of Os XPO1 in stress response and plant development.