A survey of ancient conserved non-coding elements in the PAX6 locus reveals a landscape of interdigitated cis-regulatory archipelagos.
Biological differences between cell types and developmental processes are characterised by differences in gene expression profiles. Gene-distal enhancers are key components of the regulatory networks that specify the tissue-specific expression patterns driving embryonic development and cell fate decisions, and variations in their sequences are a major contributor to genetic disease and disease susceptibility. Despite advances in the methods for discovery of putative cis-regulatory sequences, characterisation of their spatio-temporal enhancer activities in a mammalian model system remains a major bottle-neck. We employed a strategy that combines gnathostome sequence conservation with transgenic mouse and zebrafish reporter assays to survey the genomic locus of the developmental control gene PAX6 for the presence of novel cis-regulatory elements. Sequence comparison between human and the cartilaginous elephant shark (Callorhinchus milii) revealed several ancient gnathostome conserved non-coding elements (agCNEs) dispersed widely throughout the PAX6 locus, extending the range of the known PAX6 cis-regulatory landscape to contain the full upstream PAX6-RCN1 intergenic region. Our data indicates that ancient conserved regulatory sequences can be tested effectively in transgenic zebrafish even when not conserved in zebrafish themselves. The strategy also allows efficient dissection of compound regulatory regions previously assessed in transgenic mice. Remarkable overlap in expression patterns driven by sets of agCNEs indicates that PAX6 resides in a landscape of multiple tissue-specific regulatory archipelagos.
MRC Human Genetics Unit at the MRC IGMM at the University of Edinburgh, Edinburgh EH4 2XU, United Kingdom.
Shipra
S
Bhatia
MRC Human Genetics Unit at the MRC IGMM at the University of Edinburgh, Edinburgh EH4 2XU, United Kingdom.
Jack
J
Monahan
Institute of Molecular and Cell Biology, Agency for Science Technology and Research (A⁎STAR), Biopolis, Singapore 138673, Singapore.
Vydianathan
V
Ravi
MRC Human Genetics Unit at the MRC IGMM at the University of Edinburgh, Edinburgh EH4 2XU, United Kingdom.
Philippe
P
Gautier
MRC Human Genetics Unit at the MRC IGMM at the University of Edinburgh, Edinburgh EH4 2XU, United Kingdom.
Emma
E
Murdoch
Institute of Molecular and Cell Biology, Agency for Science Technology and Research (A⁎STAR), Biopolis, Singapore 138673, Singapore.
Sydney
S
Brenner
MRC Human Genetics Unit at the MRC IGMM at the University of Edinburgh, Edinburgh EH4 2XU, United Kingdom.
Veronica
V
van Heyningen
Institute of Molecular and Cell Biology, Agency for Science Technology and Research (A⁎STAR), Biopolis, Singapore 138673, Singapore.
Byrappa
B
Venkatesh
MRC Human Genetics Unit at the MRC IGMM at the University of Edinburgh, Edinburgh EH4 2XU, United Kingdom. Electronic address: Dirk-Jan.Kleinjan@igmm.ed.ac.uk.
Dirk A
DA
Kleinjan
D000818:Animals; D030801:Animals, Genetically Modified; D001483:Base Sequence; D002460:Cell Line; D002645:Chickens / Q000235:genetics; D017124:Conserved Sequence / Q000235:genetics; D004742:Enhancer Elements, Genetic / Q000235:genetics*; D005123:Eye / Q000196:embryology* / Q000378:metabolism; D005136:Eye Proteins / Q000235:genetics*; D018507:Gene Expression Regulation, Developmental; D050437:Genes, Developmental / Q000235:genetics; D018398:Homeodomain Proteins / Q000235:genetics*; D006801:Humans; D051379:Mice; D009893:Opossums / Q000235:genetics; D000071841:PAX6 Transcription Factor; D051761:Paired Box Transcription Factors / Q000235:genetics*; D011401:Promoter Regions, Genetic; D022661:RNA, Untranslated / Q000235:genetics*; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics*; D012097:Repressor Proteins / Q000235:genetics*; D017422:Sequence Analysis, DNA; D012754:Sharks / Q000235:genetics; D014714:Vertebrates / Q000235:genetics; D014981:Xenopus / Q000235:genetics; D015027:Zebrafish / Q000235:genetics
D005136:Eye Proteins; D018398:Homeodomain Proteins; D000071841:PAX6 Transcription Factor; C000605730:PAX6 protein, human; D051761:Paired Box Transcription Factors; C000605732:Pax6 protein, mouse; D022661:RNA, Untranslated; D012097:Repressor Proteins
Ancient conserved gnathostome enhancers; Elephant shark; Long-range enhancers; PAX6; Regulatory archipelagos; Reporter transgenics; cis-Regulatory landscape