Naturally Segregating Variation at Ugt86Dd Contributes to Nicotine Resistance in Drosophila melanogaster.
Naturally Segregating Variation at Ugt86Dd Contributes to Nicotine Resistance in Drosophila melanogaster.
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DOI：
10.1534/genetics.117.300058
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发表时间：
2017-09
期刊：
影响因子：
3.3
通讯作者：
Macdonald SJ
中科院分区：
文献类型：
--
作者：
Highfill CA;Tran JH;Nguyen SKT;Moldenhauer TR;Wang X;Macdonald SJ
识别复杂性状变异背后的序列多态是遗传学研究的一个关键目标，因为知道准确的致病分子事件有助于深入了解控制性状变异的途径。模式系统中复杂性状的遗传分析通常是从构建QTL图谱开始的，但通常无法识别原因序列多态。此前，我们在果蝇多亲本作图资源中定位了一系列与尼古丁抗性有关的QTL，并在这里使用了一系列功能测试来从分子水平上解析QTL。一个大效应QTL存在于一簇UDP-葡萄糖醛酸基转移酶上，利用缺失消除这些解毒基因亚组的定量互补测试显示，等位基因变异影响抗性。RNAseq结果表明，UGt86Dd基因在对尼古丁抗性较强的基因中有较高的表达，该基因的前部中肠特异RNA干扰(RNAi)降低了对尼古丁的抗性。我们在UGT86Dd中发现了一个分离的22bp移码缺失，在作图过程中考虑到Indel在很大程度上消除了QTL，这意味着这一事件解释了所定位的基因座的大部分影响。CRISPR/Cas9编辑相对耐药的基因型以在UGt86Dd中产生病变，概括了自然发生的假定功能丧失的等位基因，导致耐药性的大幅降低。尽管缺失的这种主要影响，等位基因在野生种群中似乎非常罕见，可能只解释了该性状自然变异的一小部分。尽管如此，这个假定的致病编码Indel可以成为未来外来生物解毒的机械性探索的发射台。
Identifying the sequence polymorphisms underlying complex trait variation is a key goal of genetics research, since knowing the precise causative molecular events allows insight into the pathways governing trait variation. Genetic analysis of complex traits in model systems regularly starts by constructing QTL maps, but generally fails to identify causative sequence polymorphisms. Previously we mapped a series of QTL contributing to resistance to nicotine in a Drosophila melanogaster multiparental mapping resource and here use a battery of functional tests to resolve QTL to the molecular level. One large-effect QTL resided over a cluster of UDP-glucuronosyltransferases, and quantitative complementation tests using deficiencies eliminating subsets of these detoxification genes revealed allelic variation impacting resistance. RNAseq showed that Ugt86Dd had significantly higher expression in genotypes that are more resistant to nicotine, and anterior midgut-specific RNA interference (RNAi) of this gene reduced resistance. We discovered a segregating 22-bp frameshift deletion in Ugt86Dd, and accounting for the InDel during mapping largely eliminates the QTL, implying the event explains the bulk of the effect of the mapped locus. CRISPR/Cas9 editing of a relatively resistant genotype to generate lesions in Ugt86Dd that recapitulate the naturally occurring putative loss-of-function allele, leads to a large reduction in resistance. Despite this major effect of the deletion, the allele appears to be very rare in wild-caught populations and likely explains only a small fraction of the natural variation for the trait. Nonetheless, this putatively causative coding InDel can be a launchpad for future mechanistic exploration of xenobiotic detoxification.