DNA double-strand breaks (DSBs), such as those produced by radiation and radiomimetics, are amongst the most toxic forms of cellular damage, in part because they involve extensive oxidative modifications at the break termini. Prior to completion of DSB repair, the chemically modified termini must be removed. Various DNA processing enzymes have been implicated in the processing of these dirty ends, but molecular knowledge of this process is limited. Here, we demonstrate a role for the metallo-β-lactamase fold 5'-3' exonuclease SNM1A in this vital process. Cells disrupted for SNM1A manifest increased sensitivity to radiation and radiomimetic agents and show defects in DSB damage repair. SNM1A is recruited and is retained at the sites of DSB damage via the concerted action of its three highly conserved PBZ, PIP box and UBZ interaction domains, which mediate interactions with poly-ADP-ribose chains, PCNA and the ubiquitinated form of PCNA, respectively. SNM1A can resect DNA containing oxidative lesions induced by radiation damage at break termini. The combined results reveal a crucial role for SNM1A to digest chemically modified DNA during the repair of DSBs and imply that the catalytic domain of SNM1A is an attractive target for potentiation of radiotherapy.

• MeSH
• DNA metabolismus   genetika   MeSH
• dvouřetězcové zlomy DNA * účinky záření   MeSH
• enzymy opravy DNA * metabolismus   genetika   MeSH
• exodeoxyribonukleasy * metabolismus   genetika   MeSH
• lidé MeSH
• oprava DNA * MeSH
• proliferační antigen buněčného jádra metabolismus   genetika   MeSH
• proteiny buněčného cyklu MeSH
• ubikvitinace MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• DCLRE1A protein, human MeSH Prohlížeč
• DNA MeSH
• enzymy opravy DNA * MeSH
• exodeoxyribonukleasy * MeSH
• PCNA protein, human MeSH Prohlížeč
• proliferační antigen buněčného jádra MeSH
• proteiny buněčného cyklu MeSH

BACKGROUND: 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). OBJECTIVE: We sought to correlate the functional effect of human DCLRE1C mutations on phenotypic presentation in patients with ARTEMIS deficiency. METHODS: 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. RESULTS: 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. CONCLUSIONS: 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.

• Klíčová slova
• ARTEMIS deficiency, DCLRE1C mutations, DNA repair, V(D)J recombination, nonhomologous end-joining, severe combined immunodeficiency,
• MeSH
• alely MeSH
• B-lymfocyty fyziologie   účinky záření   MeSH
• dítě MeSH
• DNA vazebné proteiny MeSH
• dospělí MeSH
• endonukleasy MeSH
• fenotyp MeSH
• heterozygot MeSH
• histony metabolismus   MeSH
• ionizující záření MeSH
• jaderné proteiny genetika   MeSH
• kojenec MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• mutace genetika   MeSH
• mutační analýza DNA MeSH
• novorozenec MeSH
• onkogenní proteiny v-abl genetika   metabolismus   MeSH
• oprava DNA genetika   MeSH
• předškolní dítě MeSH
• těžká kombinovaná imunodeficience genetika   MeSH
• tolerance záření genetika   MeSH
• transformované buněčné linie MeSH
• V(D)J rekombinace genetika   MeSH
• Check Tag
• dítě MeSH
• dospělí MeSH
• kojenec MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• novorozenec MeSH
• předškolní dítě MeSH
• Publikační typ
• časopisecké články MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• DCLRE1C protein, human MeSH Prohlížeč
• DNA vazebné proteiny MeSH
• endonukleasy MeSH
• H2AX protein, human MeSH Prohlížeč
• histony MeSH
• jaderné proteiny MeSH
• onkogenní proteiny v-abl MeSH

Herpesvirus infections, such as those induced by human cytomegalovirus (HCMV), induce specific DNA damages. DNA damages can lead to cell mutation, death, apoptosis and immune system activation. Various types of DNA damage are repaired through multiple repair pathways, such as base excision, nucleotide excision, homologous recombination and nonhomologous end joining. Changes in the activity of DNA repair proteins during viral infection can cause disturbances in the DNA repair system and change its mechanisms. This report reviews results from studies, assaying a DNA repair system in HCMV infection.

• MeSH
• cytomegalovirové infekce enzymologie   genetika   metabolismus   mikrobiologie   MeSH
• Cytomegalovirus fyziologie   MeSH
• enzymy opravy DNA genetika   metabolismus   MeSH
• lidé MeSH
• oprava DNA * MeSH
• poškození DNA MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Názvy látek
• enzymy opravy DNA MeSH

BACKGROUND: V(D)J recombination takes place during lymphocyte development to generate a large repertoire of T- and B-cell receptors. Mutations in recombination-activating gene 1 (RAG1) and RAG2 result in loss or reduction of V(D)J recombination. It is known that different mutations in RAG genes vary in residual recombinase activity and give rise to a broad spectrum of clinical phenotypes. OBJECTIVE: We sought to study the immunologic mechanisms causing the clinical spectrum of RAG deficiency. METHODS: We included 22 patients with similar RAG1 mutations (c.519delT or c.368_369delAA) resulting in N-terminal truncated RAG1 protein with residual recombination activity but presenting with different clinical phenotypes. We studied precursor B-cell development, immunoglobulin and T-cell receptor repertoire formation, receptor editing, and B- and T-cell numbers. RESULTS: Clinically, patients were divided into 3 main categories: T(-)B(-) severe combined immunodeficiency, Omenn syndrome, and combined immunodeficiency. All patients showed a block in the precursor B-cell development, low B- and T-cell numbers, normal immunoglobulin gene use, limited B- and T-cell repertoires, and slightly impaired receptor editing. CONCLUSION: This study demonstrates that similar RAG mutations can result in similar immunobiological effects but different clinical phenotypes, indicating that the level of residual recombinase activity is not the only determinant for clinical outcome. We postulate a model in which the type and moment of antigenic pressure affect the clinical phenotypes of these patients.

• Klíčová slova
• B- and T-cell receptor repertoire, RAG deficiency, V(D)J recombination, autoimmunity, immune repertoire analysis, next generation sequencing, receptor editing,
• MeSH
• B-lymfocyty imunologie   metabolismus   MeSH
• exprese genu MeSH
• fenotyp * MeSH
• genetické asociační studie * MeSH
• genotyp MeSH
• homeodoménové proteiny genetika   metabolismus   MeSH
• hypervariabilní oblasti genetika   MeSH
• kojenec MeSH
• lidé MeSH
• mutace * MeSH
• novorozenec MeSH
• počet lymfocytů MeSH
• předškolní dítě MeSH
• T-lymfocyty imunologie   metabolismus   MeSH
• těžká kombinovaná imunodeficience diagnóza   genetika   imunologie   metabolismus   MeSH
• těžké řetězce imunoglobulinů genetika   MeSH
• V(D)J rekombinace MeSH
• Check Tag
• kojenec MeSH
• lidé MeSH
• novorozenec MeSH
• předškolní dítě MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• homeodoménové proteiny MeSH
• hypervariabilní oblasti MeSH
• RAG-1 protein MeSH Prohlížeč
• těžké řetězce imunoglobulinů MeSH