CTCF is an essential, ubiquitously expressed DNA-binding protein responsible for insulator function, nuclear architecture, and transcriptional control within vertebrates. The gene CTCF was proposed to have duplicated in early mammals, giving rise to a paralogue called “brother of regulator of imprinted sites” ( BORIS or CTCFL) with DNA binding capabilities similar to CTCF, but testis-specific expression in humans and mice. CTCF and BORIS have opposite regulatory effects on human cancer-testis genes, the anti-apoptotic BAG1 gene, the insulin-like growth factor 2/H19 imprint control region ( IGF2/H19 ICR), and show mutually exclusive expression in humans and mice, suggesting that they are antagonistic epigenetic regulators. We discovered orthologues of BORIS in at least two reptilian species and found traces of its sequence in the chicken genome, implying that the duplication giving rise to BORIS occurred much earlier than previously thought. We analysed the expression of CTCF and BORIS in a range of amniotes by conventional and quantitative PCR. BORIS, as well as CTCF, was found widely expressed in monotremes (platypus) and reptiles (bearded dragon), suggesting redundancy or cooperation between these genes in a common amniote ancestor. However, we discovered that BORIS expression was gonad-specific in marsupials (tammar wallaby) and eutherians (cattle), implying that a functional change occurred in BORIS during the early evolution of therian mammals. Since therians show imprinting of IGF2 but other vertebrate taxa do not, we speculate that CTCF and BORIS evolved specialised functions along with the evolution of imprinting at this and other loci, coinciding with the restriction of BORIS expression to the germline and potential antagonism with CTCF.
Epigenetic mechanisms heritably change gene expression without altering DNA sequence. Currently, little is known about the evolution of epigenetic traits, and the genes that control them. CTCF is an essential epigenetic regulator that is expressed widely in the tissues of vertebrates and modifies the transcription of genes by altering their location within the nucleus. CTCF duplicated at some time during vertebrate evolution, giving rise to a similar gene called BORIS with expression that is limited to parts of the testes of humans and mice, but whose function is largely unknown. BORIS may contribute to the regulation of genomic imprinting, a form of epigenetic control specific to live-bearing mammals. We discovered BORIS in all mammal groups and reptiles, implying that its genesis from CTCF occurred much earlier than previously thought, preceding genomic imprinting by over 100 million years. CTCF and BORIS have not previously been found expressed together except in tumours, leading to the hypothesis that CTCF and BORIS have conflicting functions that cause cancer when allowed to overlap. We found that CTCF and BORIS are expressed alongside each other in multiple somatic and reproductive tissues of a reptile (bearded dragon) and an egg-laying mammal (platypus), but that BORIS is restricted to the gonads of live-bearing mammals (cattle and wallaby). This indicates that BORIS specialised during mammalian evolution, in concert with the evolution of genomic imprinting.