Co-evolution and molecular adaptation of the MLO-EXO70 functional module in the course of plant terrestrialization

MLO蛋白是作为钙通道的完整膜蛋白，是光合作用真核生物所特有的。在陆地植物中，它们通常在不同物种中以多种序列多样化的亚型存在，并参与必要的生理过程，如根的形态发生和向重力性、受精以及病原菌防御。胞外是一种进化上保守的多亚单位蛋白质复合体，存在于所有真核生物中。与其他类群不同的是，在陆地植物中，Exo70胞外亚单位以多种序列多样化的亚基形式存在。我们最近发现，拟南芥MLO和EXO70形成了一个功能模块，涉及这些蛋白质的物理结合，并可能调控局部分泌，例如与细胞壁相关的货物的分泌。我们假设MLO-Exo70功能模块的关键特征是在植物陆地化过程中进化的。因此，我们建议分析Brala braunii(Charophyceae)和Mesotaum endlicherianum(Zygnematophyceae)MLO-Exo70模块的共同进化和分子适应，这两个模块作为ZCC级/分支的现存代表，标志着生物从水到陆地的转变。布氏木霉和毛细管藻基因组编码MLO样蛋白(分别为7种和3种)，在大小和膜拓扑结构上与典型的陆地植物有明显不同。此外，这些基因组分别编码两个典型的Exo70亚基。利用这组来自陆地植物姊妹种的蛋白质，我们将评估最近发现的MLO的钙通道活性，它们与细胞质钙传感器钙调蛋白结合的公认能力，以及它们与特定的Exo70家族成员的异构体偏好相互作用。我们进一步打算探索MLO和Exo70基因在不同条件下在复杂的布氏梭菌菌体不同部分的表达模式，并研究这些基因在布氏梭菌种内以及更广泛地说在轮藻亚科中的自然遗传变异。总之，这些研究将揭示一幅关于MLO-Exo70功能模块在生命从水到陆地过渡过程中推测的共同进化和分子适应的全球图景。

MLO proteins are integral membrane proteins that operate as calcium channels and that are specific to photosynthetic eukaryotes. In land plants, they are typically present in multiple sequence-diversified isoforms per species and contribute to essential physiological processes such as root thigmomorphogenesis and gravitropism, fertilization, as well as pathogen defense. Exocyst is an evolutionarily conserved multisubunit protein complex found in all eukaryotes. Different from other taxa, in land plants the EXO70 exocyst subunit is present in multiple sequence-diversified isoforms. We recently discovered that Arabidopsis thaliana MLOs and EXO70s form a functional module that involves physical association of these proteins and that likely governs focal secretion, e.g. of cell wall-related cargo. We hypothesize that key features of the MLO-EXO70 functional module evolved in the course of plant terrestrialization. We, therefore, propose analyzing the co-evolution and molecular adaptation of the MLO-EXO70 module in Chara braunii (Charophyceae) and Mesotaenium endlicherianum (Zygnematophyceae), which as an extant representatives of the so-called ZCC grade/clade mark the organismal transition from water to land. The C. braunii and M. endlicherianum genomes code for MLO-like proteins (seven and three, respectively) that in part markedly differ from their canonical land plant counterparts regarding size and membrane topology. Further, these genomes encode two prototypical EXO70 subunits each. Using this set of proteins derived from sister species of land plants, we will assess the recently discovered calcium channel activity of MLOs, their well-established ability to associate with the cytoplasmic calcium sensor calmodulin, and their isoform-preferential interaction with specific EXO70 family members. We further intend to explore the expression patterns of MLO and EXO70 genes in the various parts of the complex C. braunii thallus in different conditions and to examine the natural genetic variation of these genes within the species C. braunii and, more broadly, within the Charophyceae. Together, these studies will reveal a global picture regarding the presumed co-evolution and molecular adaptation of the MLO-EXO70 functional module in the course of transition of life from water to land.