Evolutionary conservation of the circadian gene timeout in Metazoa

<i>Timeless </i>(<i>Tim</i>) is considered to function as an essential circadian clock gene in <i>Drosophila</i>. Putative homologues of the <i>Drosophila timeless </i>gene have been identified in both mice and humans. While <i>Drosophila </i>contains two paralogs, <i>timeless </i>and <i>timeout</i>, acting in clock/light entrainment and chromosome integrity/photoreception, respectively, mammals contain only one <i>Tim </i>homolog. In this paper, we study the phylogeny of the <i>timeless</i>/<i>timeout </i>family in 48 species [including 1 protozoan (<i>Guillardia theta</i>), 1 nematode (<i>Caenorhabditis elegans</i>), 8 arthropods and 38 chordates], for which whole genome data are available by using MEGA (Molecular Evolutionary Genetics Analysis). Phylogenetic Analysis by Maximum Likelihood (PAML) was used to analyze the selective pressure acting on metazoan <i>timeless</i>/<i>timeout </i>genes. Our phylogeny clearly separates insect <i>timeless </i>and <i>timeout </i>lineages and shows that non-insect animal <i>Tim </i>genes are homologs of insect <i>timeout</i>. In this study, we explored the relatively rapidly evolving <i>timeless </i>lineage that was apparently lost from most deuterostomes, including chordates, and from <i>Caenorhabditis elegans</i>. In contrast, we found that the <i>timeout </i>protein, often confusingly called “<i>timeless</i>” in the vertebrate literature, is present throughout the available animal genomes. Selection results showed that <i>timeout </i>is under weaker negative selection than <i>timeless</i>. Finally, our phylogeny of <i>timeless</i>/<i>timeout </i>showed an evolutionary conservation of the circadian clock gene <i>timeout </i>in Metazoa. This conservation is in line with its multifunctionality, being essential for embryonic development and maintenance of chromosome integrity, among others.

<i>Timeless</i>（简写为<i>Tim</i>）被认为是果蝇（<i>Drosophila</i>）体内的核心生物钟基因。果蝇<i>timeless</i>基因的推定同源物已在小鼠和人类中被鉴定得到。果蝇拥有两个旁系同源基因：<i>timeless</i>与<i>timeout</i>，二者分别参与生物钟调控与光节律重置、以及染色体完整性维持与光感受过程；而哺乳动物仅含有一个<i>Tim</i>同源基因。本研究针对48个物种的<i>timeless</i>/<i>timeout</i>基因家族开展系统发育分析，所涉物种包括1种原生动物（<i>Guillardia theta</i>）、1种线虫（<i>Caenorhabditis elegans</i>）、8种节肢动物以及38种脊索动物，所有物种均已公开全基因组数据，分析采用分子进化遗传学分析（Molecular Evolutionary Genetics Analysis，简称MEGA）软件完成。本研究同时借助最大似然法系统发育分析（Phylogenetic Analysis by Maximum Likelihood，简称PAML），对后生动物<i>timeless</i>/<i>timeout</i>基因所受的选择压力进行解析。我们构建的系统发育树清晰区分了昆虫<i>timeless</i>与<i>timeout</i>两个演化支，并证实非昆虫动物的<i>Tim</i>基因与果蝇<i>timeout</i>属于同源关系。本研究还对演化速率相对较快的<i>timeless</i>演化支进行了探究，该演化支显然在多数后口动物（包括脊索动物）以及秀丽隐杆线虫（<i>Caenorhabditis elegans</i>）中发生了丢失。与之相反，研究发现<i>timeout</i>蛋白（在脊椎动物相关文献中常被误称为「<i>timeless</i>」）广泛存在于已公布的动物基因组中。选择压力分析结果显示，<i>timeout</i>所受的负选择压力弱于<i>timeless</i>。最后，本研究构建的<i>timeless</i>/<i>timeout</i>基因家族系统发育树表明，生物钟基因<i>timeout</i>在后生动物中具有进化保守性，这一保守性与其多功能性相符——该基因不仅是胚胎发育所必需的，同时参与维持染色体完整性等多种生命过程。