Functional Analysis Of Arabidopsis Heat Stress Transcription Factors AtHsfA6αand AtHsfA9

High temperature is a main course which results in the decrease of both the quatlity andquantity of crops, and the improving the compentency of crop seeds against high temperatureis of great economic benefit and socal effect. Duing the lenthy process of evolution, the plantshas developped mutiple mechanism to adapt to the change of environment. When the plantsare exposed to high temperature, heat shock proteins (HSPs) accumulate.The expression of this kind of heat shock protein genes is regulated by a specific kind oftranscription factors, the heat sress transcription factors(Hsfs), which can bind to a conservedmotif, the heat shock element (HSE), during the transcription of HSP genes, thereby regulatesthe on and off of HSP genes and acomplishes the corresponding biological functions.In this study, the function of two heat shcok transcription factors, AtHsfA6a and AtHsfA9,in the compacity of high temperature resistance was explored. The main results are shown asfollows:(1) The endogenous expression of the heat shock transcription factors, AtHsfA6a andAtHsfA9, in wild-type Arabidopsis indicated that the two transcription factors can bothrespond to heat shock signaling.(2) The two target genes, AtHsfA6a and AtHsfA9, were both cloned and theoverexpression vectors were constructed using the seed specific promter At2S3. Transgenetictobacco lines were generated.(3) The plants overexpressing At2S3::AtHsfA6a were sensitive to high temperature stress.In normal conditions, the germination rate of wild type tobacco NC89seeds was lower thanthat of OEAtHsfA6a seeds, however, after heat shock stress of45℃for12h,15h,18h and24h,the difference was particularly evident. Thus, the heat shock transcription factors conferredthe crop seeds enhanced resistance to high temperature. Taking into account that the At2S3 seed specific promoter could drive high expression of exogenous genes in seed, these resultsprovided rationale for Germplasm improving plant genetic engineering.(4) During the germination stage, the root growth of the tobacco lines overexpressingAt2S3::AtHsfA6a and At2S3::AtHsfA9showed increased tolerance to heat shock treatmentcompared to the wild type plants. WT, At2S3::AtHsfA6a and At2S3::AtHsfA9seeds weregerminated on MS culture medium (normal conditons and45℃heat shock for12h). Theresults showed that, in normal conditions, the root lengths of the tobacco lines overexpressingAt2S3::AtHsfA6a and At2S3::AtHsfA9were reduced by32%å'Œ37%compared with WT,however, under the heat shock, the root lengths of both the transgenetic lines and WT werereduced, and the root length of the transgenetic lines were shorter than WT by30%å'Œ19%,suggesting that the root growth of the transgenetic plants exibitted tolerance to heat stress.(5) In normal conditions, the At2S3::AtHsfA6a overexpressing plants were distinctlydwarf compared with wild type plants. The cell size showed no obvious differences betweenthe two types of plants, however, the net photosynthetic rate was clearly lower inoverexpression lines than that in wild type plants. From these results, we could infer that thedevelopmental retardation of transgenetic plants was related to the reduced rate ofphotosynthesis, which might result in less nutriton materials.The function of two heat shcok transcription factors, AtHsfA6a and AtHsfA9, in thecompacity of high temperature resistance was studied. The data showed that, when exposed toheat shock stress, the expression level of heat shock transcription factors was up-regulated inthe wild type Arabidopsis, which indicated that the two heat shock transription factors wereheat resistant. The germination rate was higher in transgene lines than that in wile type plants,suggesting that the two heat shock transcription factors indeed prosessed the compacity ofhigh temperature resistance, which provided theoretical basis for increasing the resistance tohigh temperature of crops with the two heat shock transcription factors. Meanwhile, in thisstudy, the function of a seed specific promoter was validated by the specific and efficientexpression of heat shock transcription factors driven in seeds. The transgene tabaccoexhibitted enhanced resistance to high temperature than wild type in seed germination process.This results provided evidence for the regulatory role of seed specific promters in plant earlydevelopment period.