Felgentreff Kerstin, Lee Yu Nee, Frugoni Francesco, Du Likun, van der Burg Mirjam, Giliani Silvia, Tezcan Ilhan, Reisli Ismail, Mejstrikova Ester, de Villartay Jean-Pierre, Sleckman Barry P, Manis John, Notarangelo Luigi D
Division of Immunology, Boston Children's Hospital, Harvard Medical School, Boston, Mass.
Department of Immunology, Erasmus Medical Center, Rotterdam, The Netherlands.
J Allergy Clin Immunol. 2015 Jul;136(1):140-150.e7. doi: 10.1016/j.jaci.2015.03.005. Epub 2015 Apr 25.
The endonuclease ARTEMIS, which is encoded by the DCLRE1C gene, is a component of the nonhomologous end-joining pathway and participates in hairpin opening during the V(D)J recombination process and repair of a subset of DNA double-strand breaks. Patients with ARTEMIS deficiency usually present with severe combined immunodeficiency (SCID) and cellular radiosensitivity, but hypomorphic mutations can cause milder phenotypes (leaky SCID).
We sought to correlate the functional effect of human DCLRE1C mutations on phenotypic presentation in patients with ARTEMIS deficiency.
We studied the recombination and DNA repair activity of 41 human DCLRE1C mutations in Dclre1c(-/-) v-abl kinase-transformed pro-B cells retrovirally engineered with a construct that allows quantification of recombination activity by means of flow cytometry. For assessment of DNA repair efficacy, resolution of γH2AX accumulation was studied after ionizing radiation.
Low or absent activity was detected for mutations causing a typical SCID phenotype. Most of the patients with leaky SCID were compound heterozygous for 1 loss-of-function and 1 hypomorphic allele, with significant residual levels of recombination and DNA repair activity. Deletions disrupting the C-terminus result in truncated but partially functional proteins and are often associated with leaky SCID. Overexpression of hypomorphic mutants might improve the functional defect.
Correlation between the nature and location of DCLRE1C mutations, functional activity, and the clinical phenotype has been observed. Hypomorphic variants that have been reported in the general population can be disease causing if combined in trans with a loss-of-function allele. Therapeutic strategies aimed at inducing overexpression of hypomorphic alleles might be beneficial.
由DCLRE1C基因编码的核酸内切酶ARTEMIS是非同源末端连接途径的一个组成部分，在V(D)J重组过程中参与发夹结构的打开以及部分DNA双链断裂的修复。ARTEMIS缺陷患者通常表现为严重联合免疫缺陷（SCID）和细胞放射敏感性，但低表达突变可导致较轻的表型（渗漏型SCID）。
我们试图将人类DCLRE1C突变的功能效应与ARTEMIS缺陷患者的表型表现相关联。
我们研究了41种人类DCLRE1C突变在Dclre1c(-/-) v-abl激酶转化的前B细胞中的重组和DNA修复活性，这些细胞通过逆转录病毒工程构建了一个允许通过流式细胞术定量重组活性的载体。为了评估DNA修复效率，研究了电离辐射后γH2AX积累的消退情况。
对于导致典型SCID表型的突变，检测到低活性或无活性。大多数渗漏型SCID患者为1个功能丧失等位基因和1个低表达等位基因的复合杂合子，具有显著的重组和DNA修复活性残留水平。破坏C末端的缺失导致截短但部分功能的蛋白质，并且通常与渗漏型SCID相关。低表达突变体的过表达可能改善功能缺陷。
已观察到DCLRE1C突变的性质和位置、功能活性与临床表型之间的相关性。在一般人群中报道的低表达变体如果与功能丧失等位基因反式组合可能会致病。旨在诱导低表达等位基因过表达的治疗策略可能是有益的。
