School of Biological Sciences, The University of Hong Kong, Pokfulam Hong Kong, China.
Wuhan Institute of Biotechnology, Wuhan, Hubei, China.
Proteome Sci. 2014 Mar 17;12:16. doi: 10.1186/1477-5956-12-16. eCollection 2014.
Selaginella (Selaginella moellendorffii) is a lycophyte which diverged from other vascular plants approximately 410 million years ago. As the first reported non-seed vascular plant genome, Selaginella genome data allow comparative analysis of genetic changes that may be associated with land plant evolution. Proteomics investigations on this lycophyte model have not been extensively reported. Phosphorylation represents the most common post-translational modifications and it is a ubiquitous regulatory mechanism controlling the functional expression of proteins inside living organisms.
In this study, polyethylene glycol fractionation and immobilized metal ion affinity chromatography were employed to isolate phosphopeptides from wild-growing Selaginella. Using liquid chromatography-tandem mass spectrometry analysis, 1593 unique phosphopeptides spanning 1104 non-redundant phosphosites with confirmed localization on 716 phosphoproteins were identified. Analysis of the Selaginella dataset revealed features that are consistent with other plant phosphoproteomes, such as the relative proportions of phosphorylated Ser, Thr, and Tyr residues, the highest occurrence of phosphosites in the C-terminal regions of proteins, and the localization of phosphorylation events outside protein domains. In addition, a total of 97 highly conserved phosphosites in evolutionary conserved proteins were identified, indicating the conservation of phosphorylation-dependent regulatory mechanisms in phylogenetically distinct plant species. On the other hand, close examination of proteins involved in photosynthesis revealed phosphorylation events which may be unique to Selaginella evolution. Furthermore, phosphorylation motif analyses identified Pro-directed, acidic, and basic signatures which are recognized by typical protein kinases in plants. A group of Selaginella-specific phosphoproteins were found to be enriched in the Pro-directed motif class.
Our work provides the first large-scale atlas of phosphoproteins in Selaginella which occupies a unique position in the evolution of terrestrial plants. Future research into the functional roles of Selaginella-specific phosphorylation events in photosynthesis and other processes may offer insight into the molecular mechanisms leading to the distinct evolution of lycophytes.
卷柏（卷柏属）是一种石松类植物，大约在 4.1 亿年前与其他维管植物分化。作为第一个被报道的非种子维管植物基因组，卷柏基因组数据允许对可能与陆地植物进化相关的遗传变化进行比较分析。对这种石松类模型的蛋白质组学研究尚未广泛报道。磷酸化是最常见的翻译后修饰，它是一种普遍存在的调节机制，控制着生物体内蛋白质的功能表达。
在这项研究中，使用聚乙二醇分级分离和固定化金属离子亲和层析法从野生生长的卷柏中分离磷酸肽。通过液相色谱-串联质谱分析，鉴定出 1593 个独特的磷酸肽，跨越 716 个磷酸化蛋白中的 1104 个非冗余磷酸化位点，其定位得到确认。对卷柏数据集的分析揭示了与其他植物磷酸蛋白质组一致的特征，如磷酸化丝氨酸、苏氨酸和酪氨酸残基的相对比例、蛋白质 C 末端区域磷酸化位点的最高出现频率，以及蛋白结构域外磷酸化事件的定位。此外，在进化保守蛋白中还鉴定出了 97 个高度保守的磷酸化位点，表明在系统发育上不同的植物物种中，磷酸化依赖的调节机制是保守的。另一方面，对参与光合作用的蛋白质的仔细检查揭示了可能是卷柏进化特有的磷酸化事件。此外，磷酸化基序分析确定了脯氨酸导向、酸性和碱性特征，这些特征被植物中的典型蛋白激酶识别。一组卷柏特异性的磷酸化蛋白被发现富含脯氨酸导向基序类。
我们的工作提供了卷柏中第一个大规模的磷酸蛋白质图谱，它在陆地植物的进化中占据着独特的位置。对光合作用和其他过程中卷柏特异性磷酸化事件的功能作用的进一步研究，可能会深入了解导致石松类独特进化的分子机制。
