Department of Medical Genetics, Life Sciences Institute, University of British Columbia, 2350 Health Sciences Mall, Vancouver, BC, Canada, V6T 1Z3.
Epigenetics Chromatin. 2011 Jul 20;4(1):12. doi: 10.1186/1756-8935-4-12.
Endogenous retroviruses (ERVs) are parasitic sequences whose derepression is associated with cancer and genomic instability. Many ERV families are silenced in mouse embryonic stem cells (mESCs) via SETDB1-deposited trimethylated lysine 9 of histone 3 (H3K9me3), but the mechanism of H3K9me3-dependent repression remains unknown. Multiple proteins, including members of the heterochromatin protein 1 (HP1) family, bind H3K9me2/3 and are involved in transcriptional silencing in model organisms. In this work, we address the role of such H3K9me2/3 "readers" in the silencing of ERVs in mESCs.
We demonstrate that despite the reported function of HP1 proteins in H3K9me-dependent gene repression and the critical role of H3K9me3 in transcriptional silencing of class I and class II ERVs, the depletion of HP1α, HP1β and HP1γ, alone or in combination, is not sufficient for derepression of these elements in mESCs. While loss of HP1α or HP1β leads to modest defects in DNA methylation of ERVs or spreading of H4K20me3 into flanking genomic sequence, respectively, neither protein affects H3K9me3 or H4K20me3 in ERV bodies. Furthermore, using novel ERV reporter constructs targeted to a specific genomic site, we demonstrate that, relative to Setdb1, knockdown of the remaining known H3K9me3 readers expressed in mESCs, including Cdyl, Cdyl2, Cbx2, Cbx7, Mpp8, Uhrf1 and Jarid1a-c, leads to only modest proviral reactivation.
Taken together, these results reveal that each of the known H3K9me3-binding proteins is dispensable for SETDB1-mediated ERV silencing. We speculate that H3K9me3 might maintain ERVs in a silent state in mESCs by directly inhibiting deposition of active covalent histone marks.
内源性逆转录病毒（ERVs）是寄生序列，其去抑制与癌症和基因组不稳定性有关。许多 ERV 家族在小鼠胚胎干细胞（mESCs）中通过 SETDB1 沉积的组蛋白 3 赖氨酸 9 的三甲基化（H3K9me3）沉默，但 H3K9me3 依赖性抑制的机制尚不清楚。多种蛋白，包括异染色质蛋白 1（HP1）家族的成员，结合 H3K9me2/3，并参与模型生物中的转录沉默。在这项工作中，我们研究了这些 H3K9me2/3“读取器”在 mESCs 中 ERV 沉默中的作用。
我们证明，尽管 HP1 蛋白在 H3K9me 依赖性基因抑制中的作用以及 H3K9me3 在 I 类和 II 类 ERV 的转录沉默中的关键作用已有报道，但 HP1α、HP1β 和 HP1γ 的单独或组合耗竭不足以使这些元件在 mESCs 中去抑制。虽然 HP1α 或 HP1β 的缺失分别导致 ERV 的 DNA 甲基化缺陷或 H4K20me3 扩散到侧翼基因组序列，但这两种蛋白都不影响 ERV 体中的 H3K9me3 或 H4K20me3。此外，使用靶向特定基因组位点的新型 ERV 报告构建体，我们证明，与 Setdb1 相比，在 mESCs 中表达的其他已知 H3K9me3 结合蛋白的敲低，包括 Cdyl、Cdyl2、Cbx2、Cbx7、Mpp8、Uhrf1 和 Jarid1a-c，仅导致前病毒的适度激活。
综上所述，这些结果表明，已知的每个 H3K9me3 结合蛋白对于 SETDB1 介导的 ERV 沉默都是可有可无的。我们推测 H3K9me3 可能通过直接抑制活性共价组蛋白标记的沉积，使 ERVs 在 mESCs 中保持沉默状态。
