Microtubule-Associated Protein WDL5 Is Involved In Ethylene Regulation Of Arabidopsis Response To Salt Stress

As one major component of the cytoskeleton,microtubules play a pivotal role in regulating plant cell growth and cell morphogenesis.Previous studies showed that microtubule cytoskeleton also participates in the plant responses to diverse stresses.When plants are exposed to high salinity,cortical microtubules undergo the precisely regulated dynamic process of fast depolymerization and reassembly,which are vital for seedling survival under salt stress.It is reported that some certain microtubule associated proteins participate in the adaptation to salt stress by regulating microtubule dynamics in plant cells.However,the upstream regulatory pathway and the underlying mechanism in this process remain unclear.The phytohormone ethylene plays a crucial role in the regulation of diverse stress responses.Previous studies showed that exogenously applied ethylene and its biosynthetic precursor 1-aminocyclopropane-1-carboxylic acid(ACC)can significantly increase the survival rate of Arabidopsis seedlings under salt stress.Further analysis revealed that the key transcription factor in ethylene signaling pathway ETHYLENE-INSENSITIVE 3(EIN3)is accumulated to increase the salt tolerance under high salinity.These findings suggested that ethylene is a positive regulator of plant response to salt stress.However,whether microtubule cytoskeleton is involved in the biological process of ethylene signaling regulating plant salt stress response and the underlying mechanism need further study.In the present study,the characteristics of microtubule reorganization under salt stress were observed by confocal laser scanning microscope.In the presence of ethylene receptor antagonist Ag+,there was no significant difference in the salt-induced microtubule depolymerization,but microtubule reassembly was significantly suppressed compared with the process in the control plants.Similar results were observed in the ethylene-insensitive mutants ein2-5 and ein3 eill Arabidopsis seedlings.These results together demonstrated that ethylene participates in the process of microtubule reassembly under salt stress.In addition,EIN3 plays a centrol role in ethylene-enhanced microtubule reassembly under salt stress.Thus these results also implied that EIN3 mediated transcriptional regulation is of great significance in the process of ethylene regulated microtubule reassembly under salt stress.Previous studies have shown that WAVE-DAMPENED2-LIKE5(WDL5)in WAVE-DAMPENED 2(WVD2)/WVD2-LIKE(WDL)protein family is a microtubule-stabilizing protein and promotes the formation of microtubule bundles.Further investigation revealed that WDL5,which is a target gene of EIN3,is involved in the process of ethylene signaling-inhibiting hypocotyl cell elongation in dark by affecting microtubule rearrangement.In the present study,the results of quantitative real-time PCR showed that the expression of WDL5 was up-regulated under salt stress,and this process depended on ethylene signaling.The observation of plant phenotypes and microtubule dynamics under salt stress in wdl5-1 knock out mutant and WDL5 overexpressing seedlings indicated that microtubule-associated protein WDL5 is involved in the regulation of microtubules reassembly under salt stress and functions as a positive regulator of plant salt tolerance.wdl5-1 mutant was sensitive to high salinity,while the survival rate of WDL5-overexpressing seedlings was significantly higher than that of wild type.Further genetic evidences showed that WDL5 upregulation partially rescued the salt sensitivity of ein3 eill mutant seedlings,while WDL5 knockout weakened ethylene-enhanced microtubule reassembly under salt stress.These results together demonstrated that WDL5,as a downstream target of ethylene signaling,is involved in plant salt stress response through promoting the ethylene-mediated microtubule reassembly under salt stress.Based on the above results,the present study demonstrates that ethylene signaling participates in the regulation of microtubule reassembly under salt stress.Microtubule-associated protein WDL5 is a downstream effector of ethylene signaling,and promotes microtubule reassembly under salt stress,thus enhancing salt tolerance in plants seedlings.This study reveals a novel mechanism of ethylene signaling regulating plant salt stress response and uncovers a new function of microtubule-associated protein WDL5,providing new insights into the regulatory mechanisms underlying cortical microtubule reorganization in response to salt stress.