高温对小麦基因可变剪接的影响及HSFs基因不同剪接形式功能分析

Wheat is one of the most important crops in the world. Heat stress dramatically reduces wheat yield and quality. Understanding the molecular mechanisms underlying heat stress response is becoming an urgent requirement for wheat thermotolerance improvement. Alternative splicing (AS), an post-transcriptional regulatory mechanism, plays an important role in plant stress response. However, how AS participates in heat stress response is largely unknown. Recently, heat shock factors (HSFs), the key regulators in plant heat response, were found regulated by AS, but the related mechanisms are still elusive, especially in wheat. In this proposal, a whole-genome systematic analysis of heat responsive AS will be performed by using transcriptome sequencing in wheat (seedling leaves, flag leaves and seeds at filling stage). Moreover, HSFs genes with significant AS pattern shifts during heat stress will be selected as candidates for further functional analysis. The function of different HSFs splice isoforms will be evaluated in Arabidopsis transgenic lines. Taken together, our study will present global AS landscapes in response to heat stress in wheat, and shed light on the mechanisms underlying AS regulation in plant heat stress response. Additionally, functional HSFs splice isoforms will be identified as candidates for thermotolerance improvement in wheat.

小麦是我国第二大口粮作物，高温胁迫是影响小麦生产的主要限制因素之一。解析高温胁迫响应调控机制是小麦耐热性遗传改良的重要基础。可变剪接是一种重要的基因转录后调控机制，在植物抵御环境逆境的过程中起到重要作用，但目前对其在高温胁迫响应中的研究较少。热激转录因子是植物重要耐热基因，其功能也受可变剪接的调控，目前相关研究较为初步，小麦中鲜有报道。因此，本项目拟对小麦中高温胁迫诱导的可变剪接进行全基因组分析，以阐明小麦在基因剪接层面对高温胁迫的响应程度和规律，并筛选受高温诱导可变剪接的HSFs，利用转基因的方法对其不同剪接形式进行耐热性功能鉴定。本研究可以初步揭示可变剪接对小麦及植物高温胁迫响应和对HSFs功能的调控机制，为发掘和有效利用小麦耐热基因提供理论依据。

小麦是我国重要的粮食作物，高温和干旱胁迫是影响小麦生产的重要限制因素。解析植物热旱胁迫响应分子机理是小麦耐热育种的重要基础。可变剪接是一种重要的基因转录后调控机制，对基因的表达具有关键的调控作用。但目前对可变剪接如何参与植物热旱胁迫响应研究较少。本研究综合利用生物信息学和分子生物学方法，从三个方面解析了六倍体普通小麦中基因可变剪接对高温和干旱胁迫的响应规律。.首先，我们利用转录组测序分析发现小麦在转录和剪接水平上都发生了高温胁迫诱导的全基因组水平响应，但干旱胁迫主要诱导转录表达变化，剪接水平响应较少。我们一共鉴定到9312个小麦中响应高温和干旱胁迫的可变剪接基因，这些基因显著富集于非生物逆境胁迫响应途径。.然后，我们利用生物信息学方法分析了六倍体小麦ABD亚基因组之间可变剪接的差异。分析发现来自于不同亚基因组的1026个基因在高温胁迫下表现出显著的剪接模式差异，表明基因剪接的差异能够导致六倍体小麦ABD亚基因组全基因组水平上的功能分化.最后，我们重点关注了植物耐热关键基因热激转录因子（Heat Shock Factors, HSFs）。我们利用转录组数据和RT-PCR的方法发现了9个热激转录因子基因的剪接模式在高温胁迫后发生显著变化，表明可变剪接能够调控热激转录因子基因的功能，为进一步的基因功能验证奠定基础。