Expression Analysis Of TaCIPK5?TaCIPK8 Genes In Wheat And Its Interaction Relationship With TaCBLs

Wheat is one of the most important grain crops in the world.It may be faced with many Stress factors during its growth and development,such as drought?salinity?cold?low K⁺ and so on,which may influence its productivity and quality.Therefore,it is of great significance for crop improvement to study the perception?transmission and response of stress signals in wheat.Ca²⁺,as one of the second messengers,could sense environmental stress signals and change the specific concentration of Ca²⁺ to transfer the signals to its downstream sensors,which may cause specific reactions to cope with these stresses.There are mainly four types of Ca+ sensors in plants: Calmodulins?Ca M??calmodulin-like?Cm L??Calcium-dependent protein kinases?CDPKs?and Calcineurin B-like proteins?CBLs?.CBLs is a class of protein kinases resembling Calcineurin B of yeast and animal,and could interact with CBL-interacting protein kinases?CIPKs?which form the Ca²⁺-CBLs-CIPKs complex together with CBLs and Ca²⁺.The Ca²⁺-CBLs-CIPKs complex could sense and transduct stress signals.CBLs-CIPKs network plays a crucial role in the responses to environmental stresses in plants,however,little is known in wheat compared with model plants Arabidopsis and Rice.Study on molecular level contributes to the problem of drought?salinity?cold?low K⁺ and so on in wheat breeding and field production.In this study,TaCIPK5 and TaCIPK8 genes were cloned from wheat cultivar Shi4185.Systematic bioinformatics analyses were conducted with the two encoded proteins.Real-Time PCR was used to reveal the responses of the two genes to different abiotic stresses,salt?ABA and cold.Yeast two-hybrid analysis was performed to study the interaction between TaCIPK5?TaCIPK8 and TaCBLs.Main results are as follows.1?The coding sequence of TaCIPK5 gene from wheat cultivar Shi4185 is 1395 bp,and the encolde protein have 464 amino acid residues,with isoelectric point?PI?9.31;The coding sequence of TaCIPK8 is 1350 bp,and the TaCIPK8 protein have 449 amino acid residues,with isoelectric PI 7.16.2?The result of amino acid sequence homology analysis indicated that TaCIPK5 and TaCIPK8 have high similarity with proteins in the CIPKs family.The TaCIPK5 and TaCIPK8 proteins belong to the family with ser/Thr Kinsey domain,and consist of six domains,which are N-End Kinase domain?C-End Kinase domain?NAF/FISL structure?ser/Thr Kinase domain?PPI structure and corresponding activation loop respectively.3?Wheat Shi4185 root and leaf stressed with NaCl?0.2 M??cold?4??and ABA?100 u M?were gained respectively,which were used to test the expression level of TaCIPK5 and TaCIPK8 genes.The result showed that,under the stress of NaCl,the expression level of TaCIPK5 was increased until 4 h,and then decreased in leaf;in contrast,the expression level in root was increased at 24 h.The expression level of TaCIPK8 was decreased until 12 h,and then increased in leaf;the expression level in root was increased and then decreased.Under the stress of ABA,the expression level of TaCIPK5 was increased slowly until 12 h,and then decreased in leaf;the expression level in root was increased at 24 h.The expression level of TaCIPK8 was decreased until 4h,and then increased in both leaf and root.Under the stress of cold,the expression level of TaCIPK5 was increased until 4h,and then decreased in leaf;the expression level in root was increased at 24 h.The expression level of TaCIPK8 was decreased until 4 h,and then increased in both leaf and root.Thus,the responses of TaCIPK5 and TaCIPK8 gene to the above stresses are different,and the responses of one gene to different stresses are also different.4?The yeast two-hybrid test indicated that TaCIPK5 could interact with TaCBL1?Ta CBL2?Ta CBL4?Ta CBL6 and Ta CBL7,and that TaCIPK8 could interact with Ta CBL3.Cloning of TaCIPK5 and TaCIPK8 genes,the bioinformatics analyses of the proteins encoded by TaCIPK5 and TaCIPK8,expression analyses of TaCIPK5 and TaCIPK8 genes in responsed to multiple stresses,interaction analysis between TaCIPK5?TaCIPK8 and TaCBLs suggested that TaCIPK5 and TaCIPK8 genes are involved in stress responses in wheat.The results lay foudations for further studing the fuctions of TaCIPK5 and TaCIPK8 genes and CBLs-CIPKs complexes.