Gupta Aditi, Hwang Bor-Jang, Benyamien-Roufaeil Daniel, Jain Sara, Liu Sophie, Gonzales Rex, Brown Robert A, Zalzman Michal, Lu A-Lien
University of Maryland School of Medicine, Baltimore, MD, USA.
University of Maryland School of Medicine; The Center for Stem Cell Biology and Regenerative Medicine; Marlene and Stewart Greenbaum Cancer Center, Baltimore, MD 21201, USA.
OBM Geriat. 2022;6(2). doi: 10.21926/obm.geriatr.2202196. Epub 2022 Apr 2.
Telomeres consist of special features and proteins to protect the ends of each chromosome from deterioration and fusion. The telomeric DNA repeats are highly susceptible to oxidative damage that can accelerate telomere shortening and affect telomere integrity. Several DNA repair factors including MYH/MUTYH DNA glycosylase, its interacting partners Rad9/Rad1/Hus1 checkpoint clamp, and SIRT6 aging regulator, are associated with the telomeres. MYH prevents C:G to A:T mutation by removing adenine mispaired with a frequent oxidative DNA lesion, 8-oxoguanine. Here, we show that knockout (KO) human HEK-293T cells are more sensitive to HO treatment, have higher levels of DNA strand breaks and shorter telomeres than the control cells. SIRT6 foci increase at both the global genome and at telomeric regions in HO-treated cells. However, in untreated KO HEK-293T cells, SIRT6 foci only increase at the global genome, but not at the telomeric regions. In addition, the KO HEK-293T cells have increased extra-chromosomal and intra-chromosomal telomeres compared to the control cells, even in the absence of HO treatment. After HO treatment, the frequency of extra-chromosomal telomeres increased in control HEK-293T cells. Remarkably, in HO-treated KO cells, the frequencies of extra-chromosomal telomeres, intra-chromosomal telomeres, and telomere fusions are further increased. We further found that the sensitivity to HO and shortened telomeres of KO cells, are restored by expressing wild-type hMYH, and partially rescued by expressing hMYH mutant (defective in Hus1 interaction only), but not by expressing hMYH mutant (defective in both SIRT6 and Hus1 interactions). Thus, MYH interactions with SIRT6 and Hus1 are critical for maintaining cell viability and telomeric stability. Therefore, the failure to coordinate 8-oxoG repair is detrimental to telomere integrity.
端粒由特殊特征和蛋白质组成，以保护每条染色体的末端免于退化和融合。端粒DNA重复序列极易受到氧化损伤，这种损伤会加速端粒缩短并影响端粒完整性。包括MYH/MUTYH DNA糖基化酶、其相互作用伙伴Rad9/Rad1/Hus1检查点钳以及SIRT6衰老调节因子在内的几种DNA修复因子与端粒相关。MYH通过去除与常见氧化DNA损伤8-氧代鸟嘌呤错配的腺嘌呤来防止C:G到A:T的突变。在这里，我们表明，与对照细胞相比，基因敲除（KO）的人HEK-293T细胞对过氧化氢（HO）处理更敏感，具有更高水平的DNA链断裂和更短的端粒。在HO处理的细胞中，SIRT6焦点在全基因组和端粒区域均增加。然而，在未处理的KO HEK-293T细胞中，SIRT6焦点仅在全基因组中增加，而在端粒区域不增加。此外，与对照细胞相比，即使在没有HO处理的情况下，KO HEK-293T细胞的染色体外和染色体内端粒也有所增加。HO处理后，对照HEK-293T细胞中染色体外端粒的频率增加。值得注意的是，在HO处理的KO细胞中，染色体外端粒、染色体内端粒和端粒融合的频率进一步增加。我们进一步发现，通过表达野生型hMYH可以恢复KO细胞对HO的敏感性和缩短的端粒，通过表达hMYH突变体（仅在与Hus1相互作用方面有缺陷）可部分挽救，但通过表达hMYH突变体（在与SIRT6和Hus1相互作用方面均有缺陷）则不能。因此，MYH与SIRT6和Hus1的相互作用对于维持细胞活力和端粒稳定性至关重要。因此，未能协调8-氧代鸟嘌呤的修复对端粒完整性有害。
