Li Mingai, Xu Jia, Algarra Alarcon Alberto, Carlin Silvia, Barbaro Enrico, Cappellin Luca, Velikova Violeta, Vrhovsek Urska, Loreto Francesco, Varotto Claudio
Department of Biodiversity and Molecular Ecology, Research and Innovation Centre, Fondazione Edmund Mach, San Michele all'Adige (TN), Italy.
Dipartimento di Biologia, Università di Padova, Padova, Italy.
Mol Biol Evol. 2017 Oct 1;34(10):2583-2599. doi: 10.1093/molbev/msx178.
Isoprene is the most abundant biogenic volatile hydrocarbon compound naturally emitted by plants and plays a major role in atmospheric chemistry. It has been proposed that isoprene synthases (IspS) may readily evolve from other terpene synthases, but this hypothesis has not been experimentally investigated. We isolated and functionally validated in Arabidopsis the first isoprene synthase gene, AdoIspS, from a monocotyledonous species (Arundo donax L., Poaceae). Phylogenetic reconstruction indicates that AdoIspS and dicots isoprene synthases most likely originated by parallel evolution from TPS-b monoterpene synthases. Site-directed mutagenesis demonstrated invivo the functional and evolutionary relevance of the residues considered diagnostic for IspS function. One of these positions was identified by saturating mutagenesis as a major determinant of substrate specificity in AdoIspS able to cause invivo a dramatic change in total volatile emission from hemi- to monoterpenes and supporting evolution of isoprene synthases from ocimene synthases. The mechanism responsible for IspS neofunctionalization by active site size modulation by a single amino acid mutation demonstrated in this study might be general, as the very same amino acidic position is implicated in the parallel evolution of different short-chain terpene synthases from both angiosperms and gymnosperms. Based on these results, we present a model reconciling in a unified conceptual framework the apparently contrasting patterns previously observed for isoprene synthase evolution in plants. These results indicate that parallel evolution may be driven by relatively simple biophysical constraints, and illustrate the intimate molecular evolutionary links between the structural and functional bases of traits with global relevance.
异戊二烯是植物自然释放的最丰富的生物源挥发性碳氢化合物，在大气化学中起着重要作用。有人提出异戊二烯合酶（IspS）可能很容易从其他萜类合酶进化而来，但这一假设尚未得到实验研究。我们从单子叶植物（禾本科芦竹）中分离出第一个异戊二烯合酶基因AdoIspS，并在拟南芥中对其进行了功能验证。系统发育重建表明，AdoIspS和双子叶植物异戊二烯合酶很可能由TPS-b单萜合酶平行进化而来。定点诱变在体内证明了被认为是IspS功能诊断性的残基的功能和进化相关性。通过饱和诱变确定其中一个位置是AdoIspS底物特异性的主要决定因素，能够在体内导致总挥发性排放从半萜烯到单萜烯的显著变化，并支持异戊二烯合酶从罗勒烯合酶进化而来。本研究中证明的通过单个氨基酸突变调节活性位点大小实现IspS新功能化的机制可能具有普遍性，因为相同的氨基酸位置与被子植物和裸子植物不同短链萜类合酶的平行进化有关。基于这些结果，我们提出了一个模型，在一个统一的概念框架中协调先前在植物异戊二烯合酶进化中观察到的明显对比模式。这些结果表明，平行进化可能由相对简单的生物物理限制驱动，并说明了具有全球相关性的性状的结构和功能基础之间密切的分子进化联系。
