Partial MCM4 deficiency in patients with growth retardation, adrenal insufficiency, and natural killer cell deficiency
Laure Gineau, … , Jean-Laurent Casanova, Emmanuelle Jouanguy
Laure Gineau, … , Jean-Laurent Casanova, Emmanuelle Jouanguy
Published February 22, 2012
Citation Information: J Clin Invest. 2012;122(3):821-832. https://doi.org/10.1172/JCI61014.
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Research Article

Partial MCM4 deficiency in patients with growth retardation, adrenal insufficiency, and natural killer cell deficiency

Abstract

Natural killer (NK) cells are circulating cytotoxic lymphocytes that exert potent and nonredundant antiviral activity and antitumoral activity in the mouse; however, their function in host defense in humans remains unclear. Here, we investigated 6 related patients with autosomal recessive growth retardation, adrenal insufficiency, and a selective NK cell deficiency characterized by a lack of the CD56dim NK subset. Using linkage analysis and fine mapping, we identified the disease-causing gene, MCM4, which encodes a component of the MCM2-7 helicase complex required for DNA replication. A splice-site mutation in the patients produced a frameshift, but the mutation was hypomorphic due to the creation of two new translation initiation methionine codons downstream of the premature termination codon. The patients’ fibroblasts exhibited genomic instability, which was rescued by expression of WT MCM4. These data indicate that the patients’ growth retardation and adrenal insufficiency likely reflect the ubiquitous but heterogeneous impact of the MCM4 mutation in various tissues. In addition, the specific loss of the NK CD56dim subset in patients was associated with a lower rate of NK CD56bright cell proliferation, and the maturation of NK CD56bright cells toward an NK CD56dim phenotype was tightly dependent on MCM4-dependent cell division. Thus, partial MCM4 deficiency results in a genetic syndrome of growth retardation with adrenal insufficiency and selective NK deficiency.

Authors

Laure Gineau, Céline Cognet, Nihan Kara, Francis Peter Lach, Jean Dunne, Uma Veturi, Capucine Picard, Céline Trouillet, Céline Eidenschenk, Said Aoufouchi, Alexandre Alcaïs, Owen Smith, Frédéric Geissmann, Conleth Feighery, Laurent Abel, Agata Smogorzewska, Bruce Stillman, Eric Vivier, Jean-Laurent Casanova, Emmanuelle Jouanguy

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Figure 2

The MCM4 mutation is not associated with a loss of expression.

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The MCM4 mutation is not associated with a loss of expression. (A) A...
(A) Automated sequencing profiles showing the homozygous MCM4 c.70_71insG mutation in genomic DNA extracted from EBV-B cells of a patient and a WT control. Bottom: Schematic diagram of the structure of the MCM4 gene, consisting of 16 or 17 exons (Roman numerals), indicating the position of the mutation, which affects the acceptor splice site of intron 1. (B) Sequencing profile of a patient and a control indicating the insertion of a G nucleotide in the cDNA extracted from EBV-B cells. The position of the mutation is shown on a diagram of the MCM4 gene. The dotted lines represent the two mRNA transcripts produced from MCM4. The homozygous mutation leads to the insertion of an additional nucleotide between exons 1 and 2. (C) A schematic diagram of the MCM4 protein, which has an N-terminal serine/threonine-rich domain (dark gray) and a conserved MCM domain (light gray) including an ATP-binding site (black) toward its C terminus. The homozygous mutation results in a frameshift, creating a premature stop codon in exon 2. Bottom: Western blot analysis of MCM4 on total protein extracts from primary fibroblasts and SV40 fibroblasts from P1.3 and P2.1 and EBV-B cells from P1.2, controls. A polyclonal MCM4 antibody was used. (D) Complementation, by lentiviral particles, of SV40 fibroblasts from the controls and patients, with an empty pTRIP vector, an MCM4 WT vector, and the MCM4 c.71-2A→G mutation (MCM4 MUT). MCM4 was detected with a polyclonal antibody. The empty vector and non-transfected cells were used as a negative transfection control. In C and D, GAPDH was used as a loading control.