Department of Biological Sciences, University of South Carolina, Columbia, SC 29208, USA.
Department of Biological Sciences, University of South Carolina, Columbia, SC 29208, USA.
DNA Repair (Amst). 2023 Jun;126:103491. doi: 10.1016/j.dnarep.2023.103491. Epub 2023 Mar 31.
Hutchinson-Gilford Progeria Syndrome (HGPS) is a rare genetic condition characterized by features of accelerated aging and a life expectancy of about 14 years. HGPS is commonly caused by a point mutation in the LMNA gene which codes for lamin A, an essential component of the nuclear lamina. The HGPS mutation alters splicing of the LMNA transcript, leading to a truncated, farnesylated form of lamin A termed "progerin." Progerin is also produced in small amounts in healthy individuals by alternative splicing of RNA and has been implicated in normal aging. HGPS is associated with an accumulation of genomic DNA double-strand breaks (DSBs), suggesting alteration of DNA repair. DSB repair normally occurs by either homologous recombination (HR), an accurate, templated form of repair, or by nonhomologous end-joining (NHEJ), a non-templated rejoining of DNA ends that can be error-prone; however a good portion of NHEJ events occurs precisely with no alteration to joined sequences. Previously, we reported that over-expression of progerin correlated with increased NHEJ relative to HR. We now report on progerin's impact on the nature of DNA end-joining. We used a model system involving a DNA end-joining reporter substrate integrated into the genome of cultured thymidine kinase-deficient mouse fibroblasts. Some cells were engineered to express progerin. Two closely spaced DSBs were induced in the integrated substrate through expression of endonuclease I-SceI, and DSB repair events were recovered through selection for thymidine kinase function. DNA sequencing revealed that progerin expression correlated with a significant shift away from precise end-joining between the two I-SceI sites and toward imprecise end-joining. Additional experiments revealed that progerin did not reduce HR fidelity. Our work suggests that progerin suppresses interactions between complementary sequences at DNA termini, thereby shifting DSB repair toward low-fidelity DNA end-joining and perhaps contributing to accelerated and normal aging through compromised genome stability.
亨廷顿舞蹈病-吉福德早衰综合征（Hutchinson-Gilford Progeria Syndrome，HGPS）是一种罕见的遗传性疾病，其特征为衰老加速，预期寿命约为 14 年。HGPS 通常由 LMNA 基因突变引起，该基因编码核纤层蛋白 A（lamin A），是核纤层的重要组成部分。HGPS 突变改变了 LMNA 转录本的剪接，导致 lamin A 的截短、法尼基化形式，称为“progerin”。Progerin 也通过 RNA 的选择性剪接在健康个体中少量产生，并与正常衰老有关。HGPS 与基因组 DNA 双链断裂（double-strand breaks，DSBs）的积累有关，提示 DNA 修复的改变。DSB 修复通常通过同源重组（homologous recombination，HR）或非同源末端连接（nonhomologous end-joining，NHEJ）发生，前者是一种准确的、有模板的修复形式，后者是非模板的 DNA 末端连接，可能存在错误倾向；然而，很大一部分 NHEJ 事件确实发生，且连接序列没有改变。以前，我们报道过 progerin 的过表达与相对于 HR 的 NHEJ 增加相关。现在，我们报告了 progerin 对 DNA 末端连接性质的影响。我们使用了一种涉及整合到培养的胸苷激酶缺陷型小鼠成纤维细胞基因组中的 DNA 末端连接报告底物的模型系统。一些细胞被设计表达 progerin。通过表达内切酶 I-SceI 在整合的底物中诱导两个紧密间隔的 DSB，通过选择胸苷激酶功能来恢复 DSB 修复事件。DNA 测序表明，progerin 的表达与两个 I-SceI 位点之间精确末端连接的显著偏离以及非精确末端连接相关。进一步的实验表明，progerin 不会降低 HR 保真度。我们的工作表明，progerin 抑制了 DNA 末端互补序列之间的相互作用，从而将 DSB 修复转移到低保真度的 DNA 末端连接，并通过基因组稳定性受损，可能导致加速和正常衰老。
