The diversity of axon guidance (AG) receptors reflects gains in complexity of the animal nervous system during evolution. Members of the Roundabout (Robo) family of receptors interact with Slit proteins and play important roles in many developmental processes, including AG and neural crest cell migration. There are four members of the Robo gene family. However, the evolutionary history of Robo family genes remain obscure. We analyzed the distribution of Robo family members in metazoan species ranging in complexity from hydras to humans. We undertook a phylogenetic analysis in metazoans, synteny analysis, and ancestral chromosome mapping in vertebrates, and detected selection pressure and functional divergence among four mammalian Robo paralogs. Based on our analysis, we proposed that the ancestral Robo gene could have undergone a tandem duplication in the vertebrate ancestor; then one round of whole genome duplication events occurred before the divergence of ancestral lamprey and gnathostome, generating four paralogs in early vertebrates. Robo4 paralog underwent segmental loss in the following evolutionary process. Our results showed that Robo3 paralog is under more powerful purifying selection pressure compared with other three paralogs, which could correlate with its unique expression pattern and function. Furthermore, we found four sites under positive selection pressure on the Ig1‐2 domains of Robo4 that might interfere with its binding to Slits ligand. Diverge analysis at the amino acid level showed that Robo4 paralog have relatively greater functional diversifications than other Robo paralogs. This coincides with the fact that Robo4 predominantly functions in vascular endothelial cells but not the nervous system.

中文翻译：

---

轴突导向Robo受体家族基因的进化与多样性

轴突导向 (AG) 受体的多样性反映了进化过程中动物神经系统复杂性的增加。Roundabout (Robo) 受体家族的成员与 Slit 蛋白相互作用，并在许多发育过程中发挥重要作用，包括 AG 和神经嵴细胞迁移。Robo基因家族有四个成员。然而，Robo 家族基因的进化历史仍然模糊不清。我们分析了从九头蛇到人类的复杂后生动物物种中 Robo 家族成员的分布。我们对后生动物进行了系统发育分析、同线分析和脊椎动物的祖先染色体作图，并检测了四种哺乳动物 Robo 旁系同源物之间的选择压力和功能差异。根据我们的分析，我们提出祖先的 Robo 基因可能在脊椎动物祖先中经历了串联复制；然后在祖先七鳃鳗和有颚类分化之前发生了一轮全基因组复制事件，在早期脊椎动物中产生了四个旁系同源物。Robo4 paralog 在以下进化过程中经历了节段性丢失。我们的研究结果表明，与其他三个旁系同源物相比，Robo3 旁系同源物面临更强大的纯化选择压力，这可能与其独特的表达模式和功能有关。此外，我们在 Robo4 的 Ig1-2 结构域上发现了四个处于正选择压力下的位点，这些位点可能会干扰其与 Slits 配体的结合。氨基酸水平的发散分析表明，与其他 Robo 旁系同源物相比，Robo4 旁系同源物具有相对更大的功能多样化。