Molecular genetics of plant hormone signaling

植物激素信号传导的分子遗传学

• 批准号: RGPIN-2020-04298
• 负责人: McCourt, Peter
• 金额: $ 4.01万
• 依托单位: University of Toronto
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2020
• 资助国家: 加拿大
• 起止时间: 2020-01-01 至 2021-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=716959
• 关键词: Molecular; genetics; plant; hormone; signaling

Parasitic weeds threaten approximately 100 million subsistence framers in Africa. The most pernicious of these weeds, Striga hermonthica (Striga), infests 40% of the sub-Saharan farmland and is the biggest impediment to poverty alleviation on the continent. Presently, the most common method of Striga control is hand weeding, which is time consuming, labour-intensive and disproportionately carried out by women and children. Unfortunately, since much of the crop damage inflicted by Striga occurs before it surfaces, hand weeding is ineffective. Further compounding the situation is Striga seeds remain viable in the soil for up to 20 years, thus purging fields is extremely difficult with severe infestations often resulting in abandonment of the field. Breeding approaches are presently the most economically sought method of parasitic weed control, however, long-term resistance to Striga has not been attained. These challenges necessitates innovative approaches to eradicate Striga that exploit weaknesses in its lifecycle. To this end, Striga, requires a small molecule, Strigolactone (SL), to germinate. SLs emanate from host plant roots causing Striga to germinate and infest the host. Because Striga is an obligate parasite, researchers have tried to develop cheap stable SL analogs that stimulate Striga "suicide" seed germination in the absence of the host. The development of compounds that perturb Striga germination is predicated on mechanistically understanding SL receptors in this species. Over the past grant cycle, my group identified the key SL receptors in Striga and showed one, designated ShHTL7, is essential to Striga germination. In this grant cycle, we will use a combination of genetic and structural information to understand how ShHTL7 perceives SL. Our previous work shows this receptor is highly sensitive to picomolar levels of SLs in the plant (in vivo) but this sensitivity is not reflected in the test tube (in vitro). The nature of the ShHTL7 hypersensitivity is biochemically intriguing because ShHTL7 has a large ligand binding domain (LBD) that should, in principle, decrease not increase its affinity to SL. To reconcile in vivo with in vitro observations we have developed three approaches. First, we will use a "reverse genetics" approach to identify key amino acids in the LBD that contribute to SL sensitivity. Second, we will use an unbiased "forward genetics" approach to also identify key amino acids in the ShHTL7 LBD. This second approach will also identify any ShHTL7 partner proteins that may contribute to SL sensitivity. Finally, we will develop a yeast based high throughput system to assay Striga SL receptors. The basic information gleaned will not only enlighten our understanding of this uniquely hypersensitive parasitic plant SL receptor but will also direct the development of new lead compounds that perturb Striga germination. This in turn, may have long term utility in curbing Striga infestations in Africa.

寄生杂草威胁着非洲约1亿名自给自足的农民。这些杂草中最有害的一种是草本植物，它侵袭了撒哈拉以南40%的农田，是非洲大陆减贫的最大障碍。目前，最常见的控制稻瘟的方法是手工除草，这是一项耗时、劳力密集的工作，而且主要由妇女和儿童进行。不幸的是，由于稻草对作物造成的损害大多发生在它浮出水面之前，所以手工除草是无效的。使情况进一步恶化的是，稻草种子在土壤中存活长达20年，因此净化田地极其困难，严重的感染往往导致田地被遗弃。育种方法是目前寻求的最经济的寄生杂草控制方法，然而，尚未获得对稻瘟病菌的长期抗性。这些挑战要求采取创新的办法根除利用其生命周期中的弱点的Striga。为此，Striga需要一种小分子Strigolacone(SL)才能萌发。SLS从寄主植物的根中发出，导致Striga萌发并侵染寄主。由于Striga是一种专性寄生虫，研究人员试图开发出廉价稳定的SL类似物，在没有寄主的情况下刺激Striga的“自杀性”种子萌发。 扰乱Striga萌发的化合物的发展是基于对该物种中SL受体的机械理解。在过去的资助周期中，我的团队鉴定了Striga中的关键SL受体，并证明了一个名为ShHTL7的受体对Striga的萌发是必不可少的。在这个资助周期中，我们将结合遗传和结构信息来了解ShHTL7是如何看待SL的。我们之前的工作表明，这种受体对植物体内SLS的皮摩尔水平高度敏感，但这种敏感性在试管中(在体外)没有反映出来。ShHTL7超敏反应的本质在生物化学上很有趣，因为ShHTL7有一个大的配体结合结构域(LBD)，原则上应该降低而不是增加它与SL的亲和力。为了使体内观察与体外观察相一致，我们开发了三种方法。首先，我们将使用“反向遗传学”方法来确定LBD中导致SL敏感性的关键氨基酸。其次，我们将使用一种无偏见的“正向遗传学”方法来识别ShHTL7 LBD中的关键氨基酸。这第二种方法也将确定任何可能有助于SL敏感性的ShHTL7伙伴蛋白。最后，我们将开发一个基于酵母的高通量系统来检测striga SL受体。所收集的基本信息不仅将启发我们对这种独特的高度敏感的寄生植物SL受体的理解，还将指导开发干扰Striga萌发的新的先导化合物。反过来，这可能会对遏制非洲的斯特里加灾害产生长期的效用。