A small GTP-binding gene ScRan from sugarcane,is involved in responses to various hormone stresses and Sporisirium scitamineum challenge

【作者】 黄宁； Hui Ling； Xu Zhang； Huaying Mao； Yachun Su； Weihua Su； Feng Liu； Liping Xu； Rukai Chen； 阙友雄；

【Author】 Ning Huang;Hui Ling;Xu Zhang;Huaying Mao;Yachun Su;Weihua Su;Feng Liu;Liping Xu;Rukai Chen;Youxiong Que;Key Laboratory of Sugarcane Biology and Genetic Breeding,Ministry of Agriculture,Fujian Agriculture and Forestry University;

【机构】 Key Laboratory of Sugarcane Biology and Genetic Breeding,Ministry of Agriculture,Fujian Agriculture and Forestry University；

【摘要】 [Background]The ras-like nuclear GTP-binding protein（Ran） is known as a molecular switch and plays an important role in eukaryotic metabolism.A number of studies have shown that the Ran in plants was associated with external abiotic stimuli,such as light,low temperature,Mn toxicity,hormones,drought and salt stresses,however there was no report on its involvement in disease resistance;[Materials and Methods]The disease-free root,bud,leaf sheath,stem pith and epidermis from the YC05-179 plants at the mature stage were cut and used as experimental samples.The disease-free plantlets were separately treated with 100 umol/L ABA,5 mmol/L SA,100 umol/L MeJA and 5 × 10⁶ mL^-1 smut spore suspension.The ScRan protein was analyzed by bioinformatics,prokaryotic expression,transient expression.The expression patterns of ScRan gene under various hormone stresses and Sporisirium scitamineum challenge were analyzed by qRT-PCR;[Results and discussion]An 836 bp-length sugarcane Ran gene（ScRan） was cloned from sugarcane smut-resistant genotype YC05-179.Bioinformatic analysis revealed that the ScRan gene contained a 666 bp-length complete open reading frame,encoding a stable 25 kDa-acidic protein with four core domains GDGGTGKT（I）,DTAG（II）,NKVD（III） and EISAK（IV）,and without the signal peptide.Sequence alignment and phylogenetic analysis indicated that ScRan protein was conservative in evolution.ScRan protein was located in both the nucleus and the cytoplasm and could interact with sugarcane Sc 14-3-3 protein.qRT-PCR analysis demonstrated that the expression of ScRan was higher in root than all the other sugarcane tissues including bud,leaf sheath,stem pith and epidermis.The upregulation of ScRan in case of abscisic acid（ABA）,salicylic acid（SA）,and methyl jasmonic acid（MeJA） stimuli,indicated its positive involvement in sugarcane responses to phytohormone ABA,.SA,and MeJA signaling.ScRan was positively responded to S.scitamineum infection in the early stage but was inhibited in the later stage;[Conclusion]We have isolated a full-length cDNA sequence of ScRan gene from sugarcane.Multiple alignment and phylogenetic analysis indicated that ScRan gene encoded a typical small GTP-binding protein.ScRan protein was located and could interact with Sc14-3-3 in both the nucleus and the cytoplasm.qRT-PCR analysis suggested that the expression of ScRan gene was regulated by phytohormone SA,MeJA and ABA,and was positively responded to S.scitamineum infection in the early stage but was inhibited in the later stage.更多还原

【Abstract】 [Background]The ras-like nuclear GTP-binding protein（Ran） is known as a molecular switch and plays an important role in eukaryotic metabolism.A number of studies have shown that the Ran in plants was associated with external abiotic stimuli,such as light,low temperature,Mn toxicity,hormones,drought and salt stresses,however there was no report on its involvement in disease resistance;[Materials and Methods]The disease-free root,bud,leaf sheath,stem pith and epidermis from the YC05-179 plants at the mature stage were cut and used as experimental samples.The disease-free plantlets were separately treated with 100 umol/L ABA,5 mmol/L SA,100 umol/L MeJA and 5 × 10⁶ mL^-1 smut spore suspension.The ScRan protein was analyzed by bioinformatics,prokaryotic expression,transient expression.The expression patterns of ScRan gene under various hormone stresses and Sporisirium scitamineum challenge were analyzed by qRT-PCR;[Results and discussion]An 836 bp-length sugarcane Ran gene（ScRan） was cloned from sugarcane smut-resistant genotype YC05-179.Bioinformatic analysis revealed that the ScRan gene contained a 666 bp-length complete open reading frame,encoding a stable 25 kDa-acidic protein with four core domains GDGGTGKT（I）,DTAG（II）,NKVD（III） and EISAK（IV）,and without the signal peptide.Sequence alignment and phylogenetic analysis indicated that ScRan protein was conservative in evolution.ScRan protein was located in both the nucleus and the cytoplasm and could interact with sugarcane Sc 14-3-3 protein.qRT-PCR analysis demonstrated that the expression of ScRan was higher in root than all the other sugarcane tissues including bud,leaf sheath,stem pith and epidermis.The upregulation of ScRan in case of abscisic acid（ABA）,salicylic acid（SA）,and methyl jasmonic acid（MeJA） stimuli,indicated its positive involvement in sugarcane responses to phytohormone ABA,.SA,and MeJA signaling.ScRan was positively responded to S.scitamineum infection in the early stage but was inhibited in the later stage;[Conclusion]We have isolated a full-length cDNA sequence of ScRan gene from sugarcane.Multiple alignment and phylogenetic analysis indicated that ScRan gene encoded a typical small GTP-binding protein.ScRan protein was located and could interact with Sc14-3-3 in both the nucleus and the cytoplasm.qRT-PCR analysis suggested that the expression of ScRan gene was regulated by phytohormone SA,MeJA and ABA,and was positively responded to S.scitamineum infection in the early stage but was inhibited in the later stage.更多还原