TIN2 deficiency leads to ALT-associated phenotypes and differentiation defects in embryonic stem cells

Telomere integrity is critical for embryonic development and core telomere-binding proteins such as TIN2 are key to maintaining telomere stability. Here we report that homozygous Tin2S341X resulted in embryonic lethality in mice and reduced expression of Tin2 in the derived mouse embryonic stem cells (mESCs). Homozygous mutant mESCs were able to self-renew and remain undifferentiated but displayed many phenotypes associated with alternative lengthening of telomeres (ALT), including excessively long and heterogeneous telomeres, increased ALT-associated PML bodies, and unstable chromosomal ends. These cells also showed upregulation of Zscan4 expression and elevated targeting of DAXX/ATRX and H3K9me3 marks on telomeres. Furthermore, the mutant mESCs were impeded in their differentiation capacity. Upon differentiation, DAXX/ATRX and PML bodies disassociated from telomeres in these cells, where elevated DNA damage was also apparent. Our results reveal differential responses to telomere dysfunction in mESCs versus differentiated cells and highlight the critical role of TIN2 in embryonic development. RNA-seq from 4 mESC lines including WT-1, WT-2, Tin2 S341X-1, Tin2 S341X-2. Each line has three technical replicates.

端粒完整性（Telomere integrity）对于胚胎发育至关重要，而诸如TIN2（TIN2）这类核心端粒结合蛋白是维持端粒稳定性的关键。本研究发现，纯合子Tin2S341X会导致小鼠胚胎致死，并在衍生的小鼠胚胎干细胞（mouse embryonic stem cells，mESCs）中降低Tin2的表达水平。纯合突变型mESCs可自我更新并维持未分化状态，但呈现出多种与端粒替代延长（alternative lengthening of telomeres，ALT）相关的表型，包括端粒过长且异质性显著、ALT相关PML小体（PML bodies）增多以及染色体末端不稳定。这些细胞还表现出Zscan4表达上调，且端粒上DAXX/ATRX复合物的靶向富集及组蛋白H3赖氨酸9三甲基化（H3K9me3）标记水平均有所升高。此外，突变型mESCs的分化能力受到阻碍。在分化诱导过程中，此类细胞内的DAXX/ATRX复合物与PML小体均从端粒上解离，同时可见明显升高的DNA损伤水平。本研究结果揭示了mESCs与分化细胞对端粒功能障碍的差异化响应，并强调了TIN2在胚胎发育中的关键作用。本数据集包含4株mESC系的RNA测序数据，分别为WT-1、WT-2、Tin2 S341X-1及Tin2 S341X-2，每株细胞系均设置3次技术重复。