Sequencing of Pax6 loci from the elephant shark reveals a family of Pax6 genes in vertebrate genomes, forged by ancient duplications and divergences.
Sequencing of Pax6 loci from the elephant shark reveals a family of Pax6 genes in vertebrate genomes, forged by ancient duplications and divergences.
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DOI：
10.1371/journal.pgen.1003177
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发表时间：
2013
期刊：
影响因子：
4.5
通讯作者：
Kleinjan DA
中科院分区：
文献类型：
--
作者：
Ravi V;Bhatia S;Gautier P;Loosli F;Tay BH;Tay A;Murdoch E;Coutinho P;van Heyningen V;Brenner S;Venkatesh B;Kleinjan DA
Pax6是一种发育控制基因，在整个动物王国中对眼睛发育至关重要。此外，Pax6在中枢神经系统、嗅觉系统和胰腺的其他部分中发挥关键作用。在哺乳动物中，编码多种亚型的单个Pax6基因提供这些多效性功能。在这里，我们提供的证据表明，许多其他脊椎动物物种的基因组包含多个Pax6位点。我们对软骨象鲨（Callorhinchus milii）中含有Pax6的BAC进行了测序，发现了两个不同的Pax6位点。Pax6.1与哺乳动物的Pax6高度相似，而Pax6.2编码一个无配对的Pax6。使用同线性关系，我们确定了这种新的成对的Pax6.2基因在蜥蜴和青蛙，以及在斑马鱼和其他硬骨鱼的同源物。在斑马鱼中，两个全长Pax6重复先前已知，起源于鱼类特异性基因组重复（FSGD），并由于旁系同源特异性顺式元件丢失而以不同的模式表达。我们发现，硬骨鱼以外的斑马鱼也保持重复的全长Pax6基因座，但基因和调控结构域结构的差异表明，这些Pax6旁系同源起源于一个更古老的重复事件，因此被重新命名为Pax6.3。哺乳动物和象鲨Pax6.1基因座之间的序列比较突出了短距离和长距离保守的非编码元件（CNEs）的存在。功能分析证明了Pax6转录的远程增强子的古老作用。我们发现，在斑马鱼中的Pax6.2直系同源物的配对少，是专门在视网膜发育。对与小鼠NRE/P α内部启动子具有同源性的象鲨和斑马鱼Pax6.2 CNE的转基因分析显示了高度特异性的视网膜表达。最后，斑马鱼Pax6.2的吗啉代缺失导致“小眼”表型，支持视网膜发育中的作用。总之，我们的研究表明，在脊椎动物中的多效性功能的Pax6的Pax6基因，伪造古老的重复事件和独立的，谱系特异性的基因丢失的分歧家庭。Pax6是一种高度保守的转录因子，在眼、脑、胰腺和嗅觉系统发育中起关键作用。在哺乳动物中，多种Pax6亚型由单个Pax6基因编码，嵌入复杂的调控环境中。在这里，我们提供的证据，在其他脊椎动物物种中存在多个Pax6基因座。我们发现两个Pax6基因（Pax6.1和Pax6.2）存在于象鲨（软骨鱼）的基因组中。Pax6.1在基因座结构、蛋白质序列和表达模式方面与哺乳动物Pax6高度相似;而第二个基因Pax6.2编码缺乏配对结构域的蛋白质。我们确定了Pax6.2在其他脊椎动物基因组，如蜥蜴，非洲爪蟾，硬骨鱼的直系同源物，并显示它是重要的眼睛发育在斑马鱼。此外，我们的特点是第三Pax6（Pax6.3）目前只在一些硬骨鱼。系统发育分析表明，脊椎动物Pax6基因家族的进化历史是一个古老的重复，然后在不同的谱系中独立的基因丢失的结果。基因周围的顺式调控景观的序列比较导致了新的Pax6增强子的鉴定，这些增强子提供了不同Pax6家族成员之间的联系。
Pax6 is a developmental control gene essential for eye development throughout the animal kingdom. In addition, Pax6 plays key roles in other parts of the CNS, olfactory system, and pancreas. In mammals a single Pax6 gene encoding multiple isoforms delivers these pleiotropic functions. Here we provide evidence that the genomes of many other vertebrate species contain multiple Pax6 loci. We sequenced Pax6-containing BACs from the cartilaginous elephant shark (Callorhinchus milii) and found two distinct Pax6 loci. Pax6.1 is highly similar to mammalian Pax6, while Pax6.2 encodes a paired-less Pax6. Using synteny relationships, we identify homologs of this novel paired-less Pax6.2 gene in lizard and in frog, as well as in zebrafish and in other teleosts. In zebrafish two full-length Pax6 duplicates were known previously, originating from the fish-specific genome duplication (FSGD) and expressed in divergent patterns due to paralog-specific loss of cis-elements. We show that teleosts other than zebrafish also maintain duplicate full-length Pax6 loci, but differences in gene and regulatory domain structure suggest that these Pax6 paralogs originate from a more ancient duplication event and are hence renamed as Pax6.3. Sequence comparisons between mammalian and elephant shark Pax6.1 loci highlight the presence of short- and long-range conserved noncoding elements (CNEs). Functional analysis demonstrates the ancient role of long-range enhancers for Pax6 transcription. We show that the paired-less Pax6.2 ortholog in zebrafish is expressed specifically in the developing retina. Transgenic analysis of elephant shark and zebrafish Pax6.2 CNEs with homology to the mouse NRE/Pα internal promoter revealed highly specific retinal expression. Finally, morpholino depletion of zebrafish Pax6.2 resulted in a “small eye” phenotype, supporting a role in retinal development. In summary, our study reveals that the pleiotropic functions of Pax6 in vertebrates are served by a divergent family of Pax6 genes, forged by ancient duplication events and by independent, lineage-specific gene losses. Pax6 is a highly conserved transcription factor with key roles in eye, brain, pancreas, and olfactory system development. In mammals multiple Pax6 isoforms are encoded by a single Pax6 gene, embedded within a complex regulatory landscape. Here we provide evidence for the presence of multiple Pax6 loci in other vertebrate species. We show that two Pax6 genes (Pax6.1 and Pax6.2) are present in the genome of elephant shark (a cartilaginous fish). Pax6.1 is highly similar to mammalian Pax6 in terms of structure of the gene locus, protein sequence, and expression pattern; whereas the second gene, Pax6.2, codes for a protein lacking the paired domain. We identify orthologs of Pax6.2 in other vertebrate genomes, such as lizard, Xenopus, and teleost fishes, and show it is important for eye development in zebrafish. Additionally, we have characterised a third Pax6 (Pax6.3) present only in some teleost fishes. Phylogenetic analyses indicate that the evolutionary history of the Pax6 gene family in vertebrates is a result of ancient duplications followed by independent gene losses in different lineages. Sequence comparison of the cis-regulatory landscapes around the genes has led to the identification of novel Pax6 enhancers that provide a link between the diverged Pax6 family members.
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