A recurrent germline PAX5 mutation confers susceptibility to pre-B cell acute lymphoblastic leukemia

Somatic alterations of the lymphoid transcription factor gene PAX5 (also known as BSAP) are a hallmark of B cell precursor acute lymphoblastic leukemia (B-ALL)1–3, but inherited mutations of PAX5 have not previously been described. Here we report a new heterozygous germline variant, c.547G>A (p.Gly183Ser), affecting the octapeptide domain of PAX5 that was found to segregate with disease in two unrelated kindreds with autosomal dominant B-ALL. Leukemic cells from all affected individuals in both families exhibited 9p deletion, with loss of heterozygosity and retention of the mutant PAX5 allele at 9p13. Two additional sporadic ALL cases with 9p loss harbored somatic PAX5 substitutions affecting Gly183. Functional and gene expression analysis of the PAX5 mutation demonstrated that it had significantly reduced transcriptional activity. These data extend the role of PAX5 alterations in the pathogenesis of pre-B cell ALL and implicate PAX5 in a new syndrome of susceptibility to pre-B cell neoplasia

A Recessive Founder Mutation in Regulator of Telomere Elongation Helicase 1, <i>RTEL1</i>, Underlies Severe Immunodeficiency and Features of Hoyeraal Hreidarsson Syndrome

<div><p>Dyskeratosis congenita (DC) is a heterogeneous inherited bone marrow failure and cancer predisposition syndrome in which germline mutations in telomere biology genes account for approximately one-half of known families. Hoyeraal Hreidarsson syndrome (HH) is a clinically severe variant of DC in which patients also have cerebellar hypoplasia and may present with severe immunodeficiency and enteropathy. We discovered a germline autosomal recessive mutation in <i>RTEL1</i>, a helicase with critical telomeric functions, in two unrelated families of Ashkenazi Jewish (AJ) ancestry. The affected individuals in these families are homozygous for the same mutation, R1264H, which affects three isoforms of <i>RTEL1</i>. Each parent was a heterozygous carrier of one mutant allele. Patient-derived cell lines revealed evidence of telomere dysfunction, including significantly decreased telomere length, telomere length heterogeneity, and the presence of extra-chromosomal circular telomeric DNA. In addition, <i>RTEL1</i> mutant cells exhibited enhanced sensitivity to the interstrand cross-linking agent mitomycin C. The molecular data and the patterns of inheritance are consistent with a hypomorphic mutation in <i>RTEL1</i> as the underlying basis of the clinical and cellular phenotypes. This study further implicates <i>RTEL1</i> in the etiology of DC/HH and immunodeficiency, and identifies the first known homozygous autosomal recessive disease-associated mutation in <i>RTEL1</i>.</p></div

Inhibiting DNA replication blocks T-circle formation in MSK-41 <i>RTEL1^R1264H</i> cells.

<p>(A) Phi29-dependent T-circles in BJ hTERT and MSK-41. (B) Phi29-dependent T-circles in RTEL1 floxed/- MEFs ± Cre, BJ hTERT and MSK-41. (C) Phi29-dependent T-circles in BJ hTERT and MSK-41 ± aphidicolin (APD; 5 µM). (D) Dot blot of the Phi29-dependent T-circles in BJ hTERT and MSK-41 ± aphidicolin (APD; 5 µM). (E) Quantification of the fold increase in intensity of Phi29-dependent T-circles in the different cell lines subjected to the indicated treatments. Intensity mean and standard deviation were calculated over two independent experiments; statistical analysis (one-way ANOVA) was calculated with Prism (GraphPad).</p

Telomere length is altered in individuals with <i>RTEL1^R1264H</i>.

<p>(A) Primary lymphocyte telomeres in family NCI-318 were measured by flow cytometry with fluorescent <i>in situ</i> hybridization (FISH) <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003695#pgen.1003695-Alter2" target="_blank">[3]</a>. The proband is indicated by a triangle, the mother by a circle, and the father by a square. (B) Telomere FISH analysis of MSK-41 hTERT-immortalized fibroblasts revealed extreme telomere length heterogeneity. Quantitation of chromatids lacking detectable telomeric signal is shown. BJ hTERT, a normal hTERT-immortalized fibroblast line, and SaOS-2, an osteosarcoma cell line that relies on recombination-based telomere maintenance (ALT), are presented for comparison. (C) Representative metaphase spreads of MSK-41 and BJ hTERT are shown.</p

<i>RTEL1^R1264H</i> affects a putative conserved C4C4 domain.

<p>As displayed on the schematic (representing ENSP00000353332), the RTEL1 mutation is at the C-terminus of the protein, distal to the helicase domain. The affected amino acid is in a putative C4C4 domain. All eight key cysteines and R1264 are conserved in human, orangutan, cattle, and mouse sequences. Higher percent identity at a given amino acid position is indicated by a deeper purple color.</p

Clinical characteristics of families with <i>RTEL1</i> mutations.

<p>Abbreviations: DC, dyskeratosis congenita; HH, Hoyeraal Hreidarsson syndrome; BMF, bone marrow failure; IUGR, intra-uterine growth retardation; MUD HSCT, matched-unrelated donor hematopoietic stem cell transplantation; N/A, not applicable.</p

NCI-318 and MSK-41 pedigrees with <i>RTEL1</i> mutation and shared risk haplotype.

<p>NCI-318 (A) and MSK-41 (B) pedigrees are shown. Red symbols indicate affected individuals. The pink rectangles indicate the shared haplotype between the pedigrees. Each other colored rectangle indicates a unique haplotype.</p