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. 2009 Nov 23;19(4):573–583. doi: 10.1093/hmg/ddp524

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© The Author 2009. Published by Oxford University Press

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.5), which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Figure 2. — Huntingtin stimulated PRC2 during development. (A) Whole mount in situ hybridization Hoxb1, Hoxb2, Hoxb9 mRNA expression in E7.5 Hdh^ex^/4/5/Hdh^ex^/4/5 (KO) huntingtin null embryos was not properly silenced and restricted to the posterior, as observed for wild-type Hdh⁺/Hdh⁺ (WT) embryos (top). Decreased placental lactogen (PL-1) mRNA signal (purple) in bisected decidua revealed fewer giant trophoblast cells in female embryos lacking huntingtin (KO), compared with null male (KO) or wild-type (WT) embryos of either gender (bottom). Female embryos, at the indicated developmental stages (ectoplacental cone, Reichert's membrane removed), show aberrant Xp GFP-transgene reactivation in the Hdh^ex^/4/5/Hdh^ex^/4/5 (KO) extraembryonic tissue (Exem), compared with Hdh⁺/Hdh⁺ (WT) tissue, while random inactivation in the embryo proper (Em) appeared normal (right). (B) Confocal images of immunostained sections of day 4 Hdh⁺/Hdh⁺ (WT) and Hdh^ex^/4/5/Hdh^ex^/4/5 (KO) huntingtin null EBs revealed decreased histone H3K27me3 signal in the latter, whereas knock-in Hdh⁺/Hdh^Q¹¹¹ (KI) EBs, expressing huntingtin with a 111 residue polyglutamine segment exhibit increased histone H3K27me3 signal. Immunoblot of nuclear extracts and plot of histone H3K27me3/H3 band intensity ratio, normalized to wild-type (white bar), confirmed significantly (P < 0.016) decreased histone H3K27me3 in Hdh^ex^/4/5/Hdh^ex^/4/5 (KO) extracts (black bar) and significantly (P < 0.044) increased histone H3K27me3 in knock-in Hdh⁺/Hdh^Q¹¹¹ (KI) extract (grey bar), compared with wild-type (n = 4).