Carrillo Jaime, Calvete Oriol, Pintado-Berninches Laura, Manguan-García Cristina, Sevilla Navarro Julian, Arias-Salgado Elena G, Sastre Leandro, Guenechea Guillermo, López Granados Eduardo, de Villartay Jean-Pierre, Revy Patrick, Benitez Javier, Perona Rosario
Instituto Investigaciones Biomédicas (CSIC/UAM), Madrid, Spain.
Human Genetics Group, Spanish National Cancer Research Center (CNIO), Madrid, Spain.
Hum Mol Genet. 2017 May 15;26(10):1900-1914. doi: 10.1093/hmg/ddx098.
NHEJ1-patients develop severe progressive lymphocytopenia and premature aging of hematopoietic stem cells (HSCs) at a young age. Here we show a patient with a homozygous-NHEJ1 mutation identified by whole exome-sequencing that developed severe pancytopenia and bone marrow aplasia correlating with the presence of short telomeres. The mutation resulted in a truncated protein. In an attempt to identify the mechanism behind the short telomere phenotype found in the NHEJ1-patient we downregulated NHEJ1 expression in 293T and CD34+cells. This downregulation resulted in reduced telomerase activity and decreased expression of several telomerase/shelterin genes. Interestingly, cell lines derived from two other NHEJ1-deficient patients with different mutations also showed increased p21 expression, inhibition in expression of several telomerase complex genes and shortened telomeres. Decrease in expression of telomerase/shelterin genes did not occur when we inhibited expression of other NHEJ genes mutated in SCID patients: DNA-PK, Artemis or LigaseIV. Because premature aging of HSCs is observed only in NHEJ1 patients, we propose that is the result of senescence induced by decreased expression of telomerase/shelterin genes that lead to an inhibition of telomerase activity. Previous reports failed to find this connection because of the use of patient´s cells immortalized by TERT expression or recombined telomeres by ALT pathway. In summary, defective regulation of telomere biology together with defective V(D)J recombination can negatively impact on the evolution of the disease in these patients. Identification of telomere shortening is important since it may open new therapeutic interventions for these patients by treatments aimed to recover the expression of telomerase genes.
NHEJ1缺陷患者在年轻时会出现严重的进行性淋巴细胞减少和造血干细胞（HSC）过早衰老。在此，我们展示了一名通过全外显子测序鉴定出纯合NHEJ1突变的患者，该患者出现了严重的全血细胞减少和骨髓再生障碍，这与短端粒的存在相关。该突变导致了一种截短的蛋白质。为了确定在NHEJ1缺陷患者中发现的短端粒表型背后的机制，我们在293T细胞和CD34+细胞中下调了NHEJ1的表达。这种下调导致端粒酶活性降低以及几个端粒酶/端粒保护蛋白基因的表达减少。有趣的是，来自另外两名具有不同突变的NHEJ1缺陷患者的细胞系也显示p21表达增加、几个端粒酶复合体基因的表达受到抑制以及端粒缩短。当我们抑制在重症联合免疫缺陷（SCID）患者中发生突变的其他NHEJ基因（DNA-PK、Artemis或连接酶IV）的表达时，端粒酶/端粒保护蛋白基因的表达并未降低。由于仅在NHEJ1患者中观察到HSC过早衰老，我们提出这是由于端粒酶/端粒保护蛋白基因表达降低诱导的衰老导致端粒酶活性受到抑制的结果。先前的报告未能发现这种联系，因为使用了通过TERT表达永生化的患者细胞或通过替代延长端粒途径重组端粒的细胞。总之，端粒生物学的缺陷调节以及有缺陷的V(D)J重组会对这些患者疾病的发展产生负面影响。端粒缩短的鉴定很重要，因为它可能通过旨在恢复端粒酶基因表达的治疗方法为这些患者开辟新的治疗干预措施。
