Fast on the heels of a major epigenetic engineering breakthrough revealing that XIST can help treat the symptoms of Down’s syndrome, Dr. Carolyn Brown (University of British Columbia) and crew sought to find out exactly what’s going on at the molecular level with this complex lncRNA.
Their experimentation started with they hypothesis that “long ncRNAs may be serving as adapter molecules containing different protein-recognition motifs to recruit components of the gene-silencing machinery to the inactive X chromosome.” They then used “a single-copy FRT integration site to generate DOX-inducible XIST cDNA integrations” to “focus on refining the minimal XIST sequence necessary for cis-regulated silencing.” This was done “independent of the developmental signals that establish mono-allelic XIST expression in females.”
Here’s what they found:
- “Expression of the inducible full-length XIST cDNA was able to suppress expression of two nearby reporter genes as well as endogenous genes up to 3 MB from the integration site.”
- “Silencing of adjacent reporter genes can be effected by as little as 94bp of XIST, including two ‘monomers’ of the A repeat.”
- “This region includes a pair of essential palindromic sequences that are evolutionarily well-conserved and the first of these is likely to form an intra-repeat hairpin structure.”
- “Additional sequences are required for the spread of silencing to endogenous genes on the chromosome.”
The authors conclude, “The intricate set of events that ultimately lead to X-chromosome inactivation in female mammals remains a vanguard to mammalian epigenetic research. By focusing only on the ability to silence a proximal reporter we have reduced the complexity of deciphering the critical roles of XIST. We demonstrate that a mere 94bp-long sequence of repeat A can silence flanking reporter genes, but not more distal endogenous genes that are silenced by induction of the full-length XIST RNA.”
Check out the article over at Epigenetics and Chromatin, September 2013