Sequencing of Pax6 loci from the elephant shark reveals a family of Pax6 genes in vertebrate genomes, forged by ancient duplications and divergences.
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.
Institute of Molecular and Cell Biology, Agency for Science Technology and Research (ASTAR), Biopolis, Singapore, Singapore.
Vydianathan
V
Ravi
Shipra
S
Bhatia
Philippe
P
Gautier
Felix
F
Loosli
Boon-Hui
BH
Tay
Alice
A
Tay
Emma
E
Murdoch
Pedro
P
Coutinho
Veronica
V
van Heyningen
Sydney
S
Brenner
Byrappa
B
Venkatesh
Dirk A
DA
Kleinjan
D000818:Animals; D019143:Evolution, Molecular; D005136:Eye Proteins / Q000235:genetics* / Q000378:metabolism; D020440:Gene Duplication; D005786:Gene Expression Regulation; D014644:Genetic Variation; D016678:Genome; D018398:Homeodomain Proteins / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D000071841:PAX6 Transcription Factor; D051761:Paired Box Transcription Factors / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic; D012097:Repressor Proteins / Q000235:genetics* / Q000378:metabolism; D012160:Retina / Q000378:metabolism; D017422:Sequence Analysis, DNA; D012754:Sharks / Q000235:genetics*; D014714:Vertebrates / Q000235:genetics / Q000254:growth & development; D015027:Zebrafish / Q000235:genetics / Q000254:growth & development
D005136:Eye Proteins; D018398:Homeodomain Proteins; D000071841:PAX6 Transcription Factor; D051761:Paired Box Transcription Factors; C000605732:Pax6 protein, mouse; D012097:Repressor Proteins