Sagie Shira, Ellran Erika, Katzir Hagar, Shaked Rony, Yehezkel Shiran, Laevsky Ilana, Ghanayim Alaa, Geiger Dan, Tzukerman Maty, Selig Sara
Laboratory of Molecular Medicine, Rambam Health Care Campus and Rappaport Faculty of Medicine and Research Institute, Technion, Haifa 31096, Israel.
The Sohnis and Forman Families Stem Cell Center, Rappaport Faculty of Medicine and Research Institute, Technion, Haifa 31096, Israel.
Hum Mol Genet. 2014 Jul 15;23(14):3629-40. doi: 10.1093/hmg/ddu071. Epub 2014 Feb 18.
Human telomeric regions are packaged as constitutive heterochromatin, characterized by extensive subtelomeric DNA methylation and specific histone modifications. ICF (immunodeficiency, centromeric instability, facial anomalies) type I patients carry mutations in DNA methyltransferase 3B (DNMT3B) that methylates de novo repetitive sequences during early embryonic development. ICF type I patient fibroblasts display hypomethylated subtelomeres, abnormally short telomeres and premature senescence. In order to study the molecular mechanism by which the failure to de novo methylate subtelomeres results in accelerated telomere shortening, we generated induced pluripotent stem cells (iPSCs) from 3 ICF type I patients. Telomeres were elongated in ICF-iPSCs during reprogramming, and the senescence phenotype was abolished despite sustained subtelomeric hypomethylation and high TERRA levels. Fibroblast-like cells (FLs) isolated from differentiated ICF-iPSCs maintained abnormally high TERRA levels, and telomeres in these cells shortened at an accelerated rate, leading to early senescence, thus recapitulating the telomeric phenotype of the parental fibroblasts. These findings demonstrate that the abnormal telomere phenotype associated with subtelomeric hypomethylation is overridden in cells expressing telomerase, therefore excluding telomerase inhibition by TERRA as a central mechanism responsible for telomere shortening in ICF syndrome. The data in the current study lend support to the use of ICF-iPSCs for modeling of phenotypic and molecular defects in ICF syndrome and for unraveling the mechanism whereby subtelomeric hypomethylation is linked to accelerated telomeric loss in this syndrome.
人类端粒区域被包装成组成型异染色质，其特征是广泛的亚端粒DNA甲基化和特定的组蛋白修饰。I型ICF（免疫缺陷、着丝粒不稳定、面部异常）患者携带DNA甲基转移酶3B（DNMT3B）的突变，该酶在早期胚胎发育过程中对新生重复序列进行甲基化。I型ICF患者成纤维细胞显示亚端粒低甲基化、端粒异常缩短和早衰。为了研究亚端粒新生甲基化失败导致端粒加速缩短的分子机制，我们从3名I型ICF患者中生成了诱导多能干细胞（iPSC）。在重编程过程中，ICF-iPSC中的端粒延长，尽管亚端粒持续低甲基化和TERRA水平较高，但衰老表型被消除。从分化的ICF-iPSC中分离出的成纤维样细胞（FL）维持异常高的TERRA水平，这些细胞中的端粒以加速速率缩短，导致早衰，从而重现了亲代成纤维细胞的端粒表型。这些发现表明，与亚端粒低甲基化相关的异常端粒表型在表达端粒酶的细胞中被克服，因此排除了TERRA抑制端粒酶作为ICF综合征中端粒缩短的核心机制。本研究中的数据支持使用ICF-iPSC对ICF综合征中的表型和分子缺陷进行建模，并阐明亚端粒低甲基化与该综合征中端粒加速丢失相关的机制。
