Mechanism Studies On Cold Acclimation And Photoprotection In Evergreen Members Of Genus Rhododendron

Rhododendron is one of the most diverse genera of plants on earth,which includes both deciduous and evergreen species.Subgenus Azaleastrum,Pentanthera and Tsutsusi comprise what gardeners loosely defined as’azaleas’.The species and cultivars of subgenus Tsutsusi are usually referred as’evergreen azaleas’.In China,they are mainly located in the south of Yangzi River,where the mean minimum air temperature in winter is around 0℃.The low winter temperature is one of the critical components limiting their geographic distribution.Subgenus Rhododendron and Hymenanthes comprise what gardeners loosely refer as’evergreen rhododendrons’.In China,evergreen rhododendrons are usually located in rugged mountains with relatively low air temperatures.Though they are relatively cold-hardy,the high light intensity in winter is a stress for their survival.Photoprotective mechanisms in evergreen rhododendrons during winter time has not been explored in depth.Exploring the mechanism of cold acclimation in rhododendrons and azaleas is good for their resistant breeding and wide landscape application.In this study,1)we compared the freezing tolerance of 10 evergreen azaleas and their de-acclimated kinetics;2)One of the 10 evergreen azaleas—’Elsie Lee’with its winter LT50 lower than-20 ℃ was chosen to explore the mechanism of cold acclimation via comparing the physiological and transcriptional changes between natural and chamber cold acclimation;3)Based on the results of previous studies,photosynthetic/photoprotective response during cold acclimation was of more value for further study.Since evergreen rhododendrons are more suitable for photoprotective study,two elevational ecotypes of R.catawbiense were the plant materials for the study of this part.The main results are as follows:1.Comparative physiology of natural deacclimation in 10 evergreen azalea cultivarsTen evergreen azalea cultivars-’Changchunerhao’’Dazhusha’’Elsie Lee’’Hongshanhu’’Nuccio’s Pink Bobble’’Shiyandujuan’’Yudaizhirong’’Zhuangyuanhong’’Zihudie’’Zi’e’are as materials to explore their leaf freezing tolerance and de-acclimated characteristics from January to March in Hangzhou,China.We found:1)the winter-hardiness in 10 evergreen azalea cultivars are different.’Elsie Lee’was the most-hardy cultivar among them.2)Eight of the 10 cultivars arrived at their hardiest level on Jan.22 and first showed deacclimation when daily mean temperature over a 2-week period preceding the leaf freezing tolerance measurement was～9.5℃.Deacclimation for other two cultivars was somewhat delayed and might have involved deacclimation-reacclimation cycling before eventual deacclimation;4)’More-hardy’cultivars(’Elsie Lee’’Nuccio’s Pink Bobble’’Yudaizhirong’’Changchunerhao’and’Shiyandujuan’)deacclimated slower than the’less-hardy’ ones(’Hongshanhu’,’Dazhusha’’Zhuangyuanhong’’Zihudie,and ’Zi’e’)over the first half of the deacclimation period,and this trend reversed during the second half of the deacclimation period;4)’More-hardy’ cultivars generally had higher total soluble sugars than’less hardy’ ones at acclimated state;total soluble sugars declined during deacclimation in all cultivars and the loss was positively correlated with the loss in leaf freezing tolerance.Leaf starch content generally followed the opposite trend to that of the total soluble sugars.2.The comparative physiology between natural and chamber cold acclimation in Rhododendron ’Elsie Lee’To investigate how Rhododendron ’Elsie Lee’ obtain its leaf freezing tolerance during cold acclimation,we compared the physiological changes between natural and chamber cold acclimation and found:1)The cold acclimation is a two-stage process in Rhododendron’Elsie Lee’.The first stage was to respond to the shortened photoperiod in autumn.Though there was only a slight increase in leaf freezing tolerance in this stage,the decreased water content indicates that plants gradually stopped growth.The second stage was to respond to the low temperature in winter.The leaf freezing tolerance increased a lot during this stage,along with the accumulation of leaf sugar and anthocyanin concentrations,and exhibited various photoprotective strategies;2)Leaf tissues could obtain some of the leaf freezing tolerance through low temperature with long photoperiod,but the capacity was only half of the capacities of natural cold acclimated leaves;3)Accumulation of anthocyanin is not a prerequisite for leaf freezing tolerance.3.The comparative transcriptome profiles between natural and chamber cold acclimation in Rhododendron ’Elsie Lee’To deeply understand how Rhododendron’Elsie Lee’ obtain the leaf freezing tolerance during cold acclimation,we compared the transcriptome profiles and qPCR data between natural and chamber cold acclimation.We found:1)Eight KEGG pathways are particularly enriched in natural cold acclimation,and they are related to photosynthesis,photoprotection,lipoxygenase(LOX)pathway,and enzymic antioxidants accumulation.Eleven KEGG pathways are particularly enriched in chamber cold acclimation,and they are related to the plant respiration,i.e.,glycolysis,citric acid cycle,and amino acid metabolisms(the intermediates of glycolysis and citric acid cycle are presecurors of amnio acids).2)In the carbon metabolism,most of the differential expressed genes were up-regulated during natural cold acclimation,but most of the differential expressed genes were down-regulated during chamber cold acclimation.3)The transcription factor HY5 related to circadian rhythm and LOX related to the unsaturated fatty acid metabolism may play essential roles for increasing the winter-hardiness during cold acclimation.4)Anthocyanin accumulation may be post-transcriptional control.4.The photoprotective strategies during natural cold acclimation in R.catawbienseIn this study,we chose a North American evergreen Rhododendron species-R.catawbiense with two elevational ecotypes maintained in a common garden of Iowa State University,U.S.A.,to explore the photosynthetic and photoprotective responses during natural cold acclimation and how microclimates of the different provenances affect the plants’ adaptations to winter stresses(low temperatures together with light stress).In this study,we found:1)High elevational ecotype(1800 m)attained overall greater leaf freezing tolerance in winter while low elevational ecotype(85 m)was more responsive to short-days for the first stage of cold acclimation;2)both elevational ecotypes employed various photoprotective strategies with different degrees to survive winter.In autumn,only high elevation ecotype exhibited slight damage to the reaction centers;in winter,both ecotypes were photo-inhibited and employed primarily sustained de-epoxidation for photoprotection.From summer to winter,low elevation ecotype still used reversible xanthophyll cycle more effectively than high elevation ecotype;3)Microclimates of R.catawbiense ecotypes inform a differential physiological response to winter cold and light stresses.