Rienstra Juriaan, Carrillo-Carrasco Vanessa Polet, de Roij Martijn, Hernandez-Garcia Jorge, Weijers Dolf
Laboratory of Biochemistry, Wageningen University, Wageningen 6708WE, The Netherlands.
Proc Natl Acad Sci U S A. 2025 Apr 8;122(14):e2501915122. doi: 10.1073/pnas.2501915122. Epub 2025 Apr 1.
Auxin Response Factor (ARF) plant transcription factors are the key effectors in auxin signaling. Their DNA-Binding Domain (DBD) contains a B3 domain that allows base-specific interactions with Auxin Response Elements (AuxREs) in DNA target sites. Land plants encode three phylogenetically distinct ARF classes: the closely related A- and B-classes have overlapping DNA binding properties, contrasting with the different DNA-binding properties of the divergent C-class ARFs. ARF DNA-binding divergence likely occurred early in the evolution of the gene family, but the molecular determinants underlying it remain unclear. Here, we show that the B3 DNA-binding residues are deeply conserved in ARFs, and variability within these is only present in tracheophytes, correlating with greatly expanded ARF families. Using the liverwort , we confirm the essential role of conserved DNA-contacting residues for ARF function. We further show that ARF B3-AuxRE interfaces are not mutation-tolerant, suggesting low evolvability that has led to the conservation of the B3-DNA interface between ARF classes. Our data support the almost complete interchangeability between A/B-class ARF B3 by performing interspecies domain swaps in , even between ARF lineages that diverged over half a billion years ago. Our analysis further suggests that C-class ARF DNA-binding specificity diverged early during ARF evolution in a common streptophyte ancestor, followed by strong selection in A and B-class ARFs as part of a competition-based auxin response system.
生长素响应因子（ARF）植物转录因子是生长素信号传导中的关键效应器。它们的DNA结合结构域（DBD）包含一个B3结构域，该结构域允许与DNA靶位点中的生长素响应元件（AuxREs）进行碱基特异性相互作用。陆地植物编码三种系统发育上不同的ARF类别：密切相关的A类和B类具有重叠的DNA结合特性，这与不同的C类ARF的DNA结合特性形成对比。ARF DNA结合的差异可能在基因家族进化的早期就已发生，但其潜在的分子决定因素仍不清楚。在这里，我们表明B3 DNA结合残基在ARF中高度保守，并且这些残基内的变异性仅存在于维管植物中，这与大大扩展的ARF家族相关。利用地钱，我们证实了保守的DNA接触残基对ARF功能的重要作用。我们进一步表明，ARF B3-AuxRE界面不耐突变，这表明其进化能力较低，导致了ARF类别之间B3-DNA界面的保守性。我们的数据通过在 中进行种间结构域交换，支持了A/B类ARF B3之间几乎完全的互换性，甚至在超过5亿年前分化的ARF谱系之间也是如此。我们的分析进一步表明，C类ARF DNA结合特异性在共同的链形植物祖先的ARF进化早期就已分化，随后在A类和B类ARF中受到强烈选择，作为基于竞争的生长素响应系统的一部分。
