胡杨干旱响应转录组及NF-YB基因表达谱
摘要
本文研究了来自中国乌鲁木齐半干旱地区的胡杨材料对干旱适应性响应的转录水平上的基因表达及NF-YB基因在干旱响应中的表达行为。通过设置4组7周控水产生的逐渐增强的干旱胁迫水平,即从最弱强度的干旱组1到最严重干旱的干旱组4,用Affymetrix杨树基因组芯片比较分析了不同干旱强度下的胡杨转录表达谱变化。随着干旱严重度在干旱组1、干旱组2、干旱组3和干旱组4之间的增加,胡杨上调或下调转录表达水平改变的基因数量也随之增加,分别是952个、1354个、2138和2360；仅有277个基因在所有4个干旱处理中都表现出一致的表达变化；有1938个表达转录子表现出干旱强度的专化性。在表达丰度上有显著变化的基因来自AP2/EREPB, bZIP,NAC,NF-Y,WRKY,MYB和Homeobox等转录因子家族的成员；以及小热激蛋白、HSP70和HSP90等家族中的成员。转录表达数据分析说明胡杨能根据感受的不同干旱程度激活相应的响应调节途径。这些结果为了解杨树干旱胁迫响应的分子机制以及未来实验的方向提供了重要的参考。同时,为阐明木本植物中核转录Y因子B亚基蛋白(NF-YB)及其编码基因在抗旱转录调控中的作用,运用生物信息学知识和方法,从杨树模式种毛果杨基因组中鉴别出了20个PtNF-YB基因,并利用拟南芥同源基因进行了功能注释,这些基因的进化亲缘关系和同源性预测研究表明,杨树比较起拟南芥在一些未知功能组内扩增了成员。并以PtNF-YB序列为参考,从胡杨叶组织基因组材料中反转录获取了核酸长度为543bp的目标基因PeNF-YB7,GFP亚细胞定位表明该基因在核中进行转录表达功能。实时荧光定量PCR分析PEG-6000处理的胡杨材料,发现胡杨叶组织中20个NF-YB同源基因均有表达变化,至少有4个成员在干旱渗透胁迫中是表达上调。本研究首次系统地从杨树基因组中确定全部NF-YB家族基因序列、以及开展了与草本拟南芥NF-YB基因的亲缘遗传关系、同源性预测以及干旱胁迫响应表达谱的研究。研究结果不仅说明了NF-YB转录因子在木本植物中的抗旱作用,也为树木抗旱调控机制的阐明和木本植物NF-YB转录因子及相关研究起到了促进作用和提供了参考。
Populus euphratica is native to semi-arid regions of the Xinjiang Uyghur Autonomous Region of China, and studying its drought responses will greatly increase the understanding of how trees acclimate to drought. Water was withheld for seven weeks in four different drought stress treatments, with regime1being the least drought stressed and regime4being the most, and the poplar's transcriptional profiles examined with Affymetrix Poplar GeneChip microarrays. The number of significantly up or down transcriptional changes increased with the severity of drought stress, with regime1,2,3and4showing952,1354,2138and2360altered transcripts, respectively. Only277of these were found in common across all four regimes, while1938transcripts were found to be unique to the individual treatments. Genes with altered transcript abundance included members of the transcription factor families AP2/EREPB, bZIP, NAC, NF-Y, WRKY, MYB and Homeobox, as well as genes for the small HSP, HSP70and HSP90heat shock protein families. Analysis of the transcript data from these experiments indicated that P. euphratica activates specific regulatory pathways according to the degree of drought stress it receives. These results provide important insights into the molecular mechanisms underpinning the drought stress responses of poplar, as well as providing candidates for future experimentation. In addition, to identify and characterize the NF-YB genes in the model tree Populus trichocarpa, with the expectation of determining which orthologs in P. euphratica, were involved in drought response, a total of20NF-YB genes in the P. trichocarpa genome were identified and annotated by bioinformatics procedures. The examination on their phylogenetic relationships and ortholog predictions showed that Populus has an expanded set of NF-YB genes within unknown functional groups in comparison to Arabidopsis. A target sequence with543bp in length, named PeNF-YB7, from leave issues of Populus euphractica via reverse transcription approach, with the NF-YB homologue sequence in the genome of P. trichocarpa as reference. The gene was observed to be expressed in unclear field with GFP subcellular localization method. Four members of the PeNF-YB gene family in P. euphratica were observed to be up-regulated in expression during PEG-6000(polyethylene glycol6000) drought treatment based on RT-qPCR analyses. The results of this study could not only verify the role of20NF-YB genes in poplar resistance to drought, but also provide a basic clue for elucidating regulation mechanism of tree resistance to drought. The increased knowledge on the phylogenetic relationships, predicted orthologs, and expression patterns of the poplar NFYB genes during drought stress can be expected to promote the future study of woody plants.
Populus euphratica is native to semi-arid regions of the Xinjiang Uyghur Autonomous Region of China, and studying its drought responses will greatly increase the understanding of how trees acclimate to drought. Water was withheld for seven weeks in four different drought stress treatments, with regime1being the least drought stressed and regime4being the most, and the poplar's transcriptional profiles examined with Affymetrix Poplar GeneChip microarrays. The number of significantly up or down transcriptional changes increased with the severity of drought stress, with regime1,2,3and4showing952,1354,2138and2360altered transcripts, respectively. Only277of these were found in common across all four regimes, while1938transcripts were found to be unique to the individual treatments. Genes with altered transcript abundance included members of the transcription factor families AP2/EREPB, bZIP, NAC, NF-Y, WRKY, MYB and Homeobox, as well as genes for the small HSP, HSP70and HSP90heat shock protein families. Analysis of the transcript data from these experiments indicated that P. euphratica activates specific regulatory pathways according to the degree of drought stress it receives. These results provide important insights into the molecular mechanisms underpinning the drought stress responses of poplar, as well as providing candidates for future experimentation. In addition, to identify and characterize the NF-YB genes in the model tree Populus trichocarpa, with the expectation of determining which orthologs in P. euphratica, were involved in drought response, a total of20NF-YB genes in the P. trichocarpa genome were identified and annotated by bioinformatics procedures. The examination on their phylogenetic relationships and ortholog predictions showed that Populus has an expanded set of NF-YB genes within unknown functional groups in comparison to Arabidopsis. A target sequence with543bp in length, named PeNF-YB7, from leave issues of Populus euphractica via reverse transcription approach, with the NF-YB homologue sequence in the genome of P. trichocarpa as reference. The gene was observed to be expressed in unclear field with GFP subcellular localization method. Four members of the PeNF-YB gene family in P. euphratica were observed to be up-regulated in expression during PEG-6000(polyethylene glycol6000) drought treatment based on RT-qPCR analyses. The results of this study could not only verify the role of20NF-YB genes in poplar resistance to drought, but also provide a basic clue for elucidating regulation mechanism of tree resistance to drought. The increased knowledge on the phylogenetic relationships, predicted orthologs, and expression patterns of the poplar NFYB genes during drought stress can be expected to promote the future study of woody plants.
引文
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