Studies On Expression And Functions Of Rice CDPKs Genes

CDPKs take important roles in calcium-mediated signal transduction involved in plant growth and development, defenses to pathogen, responses to abiotic stresses, ionic transportation in channels. In rice, functions of most CDPKs genes are elusive with the exception of several CDPKs genes that have been known to take roles in development and responses to stresses. In the research, we identified firstly the cis-elements in responsive to stresses in promoter regions of rice CDPKs genes, then analyzed the transcription expression of CDPKs genes under different stress conditions and in different rice tissues according to the data from rice expression database (RED: http://red.dna.affrc.go.jp/RED/) and the results of RT-PCR and Northern blot. Based on expression analysis, CDPKs genes were classified and stress-responsive CDPKs genes were identified. Lastly, the functions of 0sCPK6 and 0sCPK8 were analyzed with RNAi technology and gene overexpression technology.The main results of this research are as follows:1 Transcription expression analysis of rice CDPKs genes1.1 29 rice CDPKs genes were identified in rice genome through sequence comparing with conserved domain of plant CDPKs genes; the putative amino sequences have typical CDPKs domains of Ser/Thr kinase domain and CaM-like domain.1.2 The promoter regions of CDPKs genes contain stress-responsive cis-elements of EBOXBNNAPA, IBOXCORE, MYBCORE, MYCATRD22, DRECRTCOREAT, LTREC0REATC0R15, CCAATBOX and WB0XATNPR1, which suggested that rice CDPKs genes might be in responsive to stress stimuli.1.3 11 ESTs corresponding to 11 rice CDPKs genes in rice expression database were widely in responsive to the stresses of cold, drought, salt, chitin and infection of rice blast pathogen.1.4 CDPKs genes were classified as 7 groups on the base of tissue-specific expression of CDPKs genes, and 3 CDPKs genes of 0sCPK6, OsCPK17 and OsCPK25 were firstly identified to respond to cold, drought, salt stresses and heat shock.2 The function analysis of 0sCPK82.1 RNAi technology was performed to inhibit the expression of 0sCPK8, transgenic rice plants with down-regulated expression of 0sCPK8 were obtained, the plant height of transgenic plants were shorter than that of the wild type.2.2 The plant height of transgenic plants co-segregated with transcription levels of OsCPK8 in transgenic plants, dwarf plants had low transcriptions of 0sCPK8. This result indicated that the down-regulated expression of 0sCPK8 resulted in the shortened culms of transgenic plants.2.3 The GA₃ contents in dwarf transgenic plants were lower that in WT; spraying GA₃ on dwarf plants could partially recover plant height of dwarf plants but not to the height of WT; the seedlings of dwarf plants were less sensitive to exogenous GA₃ than WT. These results indicated that the synthesis of GA and GA signal pathway were inhibited to some extent.2.4 Based on the changes of transcription of OsGA2ox1, OsGA2ox2 and OsGAI resulted from the down-regulated transcription of 0sCPK8 in dwarf plants, a putative signal pathway of OsCPK8 was presented in this paper.3 The function analysis of 0sCPK63.1 The full-length cDNA of OsCPK6 was isolated and sequenced, and the putative amino sequence had a Ser/Thr kinase domain and an EF-hand domain. OsCPK6 was located in cell membrane and nucleus.3.2 OsCPK6 was transformed into rice variety, Wuyujing 3, and 16 transgenic rice plants with single copy of exogenous gene were obtained, OsCPK6 had increased transcription in transgenic rice plants.3.3 Three transgenic rice lines of S1, S3 and S5 showed that the tolerance to drought and salt stresses increased and the rates of wilted seedling under stresses of drought and salt were lower than non-transgenic rice.3.4 Transgenic rice had higher content of proline in leaves under drought and salt stresses than negative transgenic controls. This may be physiological mechanism of increased tolerance to environmental stresses of OsCPK6 transgenic rice.3.5 The transcription of the stress-inducible transcription element gene SNAC1 was up-regulated by OsCPK6 overexpression in transgenic rice. OsCPK6 may participate in responses of rice to environmental stresses through regulating SNAC1.