Deleterious mutations in LRBA are associated with a syndrome of immune deficiency and autoimmunity - PubMed (original) (raw)

Case Reports

. 2012 Jun 8;90(6):986-1001.

doi: 10.1016/j.ajhg.2012.04.015. Epub 2012 May 17.

Giacomo Tampella, Qiang Pan-Hammarström, Peer Herholz, Claudia M Trujillo-Vargas, Kanchan Phadwal, Anna Katharina Simon, Michel Moutschen, Amos Etzioni, Adi Mory, Izhak Srugo, Doron Melamed, Kjell Hultenby, Chonghai Liu, Manuela Baronio, Massimiliano Vitali, Pierre Philippet, Vinciane Dideberg, Asghar Aghamohammadi, Nima Rezaei, Victoria Enright, Likun Du, Ulrich Salzer, Hermann Eibel, Dietmar Pfeifer, Hendrik Veelken, Hans Stauss, Vassilios Lougaris, Alessandro Plebani, E Michael Gertz, Alejandro A Schäffer, Lennart Hammarström, Bodo Grimbacher

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Case Reports

Deleterious mutations in LRBA are associated with a syndrome of immune deficiency and autoimmunity

Gabriela Lopez-Herrera et al. Am J Hum Genet. 2012.

Abstract

Most autosomal genetic causes of childhood-onset hypogammaglobulinemia are currently not well understood. Most affected individuals are simplex cases, but both autosomal-dominant and autosomal-recessive inheritance have been described. We performed genetic linkage analysis in consanguineous families affected by hypogammaglobulinemia. Four consanguineous families with childhood-onset humoral immune deficiency and features of autoimmunity shared genotype evidence for a linkage interval on chromosome 4q. Sequencing of positional candidate genes revealed that in each family, affected individuals had a distinct homozygous mutation in LRBA (lipopolysaccharide responsive beige-like anchor protein). All LRBA mutations segregated with the disease because homozygous individuals showed hypogammaglobulinemia and autoimmunity, whereas heterozygous individuals were healthy. These mutations were absent in healthy controls. Individuals with homozygous LRBA mutations had no LRBA, had disturbed B cell development, defective in vitro B cell activation, plasmablast formation, and immunoglobulin secretion, and had low proliferative responses. We conclude that mutations in LRBA cause an immune deficiency characterized by defects in B cell activation and autophagy and by susceptibility to apoptosis, all of which are associated with a clinical phenotype of hypogammaglobulinemia and autoimmunity.

Copyright © 2012 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

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Figures

Figure 1

Figure 1

Mutations in LRBA Shaded figures are affected individuals homozygous for a mutation in LRBA. Half-shaded figures are unaffected individuals heterozygous for a mutation in LRBA. (A) Family A with two affected individuals showing a homozygous missense p.Ile2657Ser substitution. (B) Family B with one affected individual who had homozygous nonsense substitution p.Arg1683∗. (C) In family C, the amino acid substitution was nonsense: p.Glu59∗. (D) The affected individual from family D has a large homozygous deletion including exons 1 and 2 and ∼60 kb upstream and ∼50 kb downstream. The parents in family D are heterozygous for the deletion, but the unaffected sibling does not carry the deletion.

Figure 2

Figure 2

Homozygous LRBA Mutations Lead to Lack of Protein (A) A schematic representation of LRBA shows domains and location of mutations at the protein level. (B) Protein extracts of Epstein-Barr virus (EBV)-transformed B cell lines (EBV cells) from individuals P1, P2, and P3, from three heterozygous members of family A, and from two controls were loaded into a polyacrylamide gel. EBV cells from homozygous affected individuals show an absence of LRBA, whereas both heterozygous individuals and the healthy controls show protein expression. (C) EBV cell lines from two affected individuals and three healthy donors were subjected to starvation. The proportion of early apoptotic cells was determined with Annexin V and propidium iodide staining. Both the percentage of early apoptotic EBV cells after starvation and the increase in apoptosis after starvation were significant (p values are 0.0172 and 0.0057, respectively, as determined by Welch t tests). We calculated the increase in apoptosis by subtracting the amount of unstarved EBV cells from the amount of starved EBV cells for both affected individuals and controls.

Figure 3

Figure 3

Phenotypic Characterization of B Cells PBMCs were gated for CD19+ CD20+ B cells (A) and were analyzed for IgM, CD27 (B), and BAFF-R expression (C), for CD10 and CD38 expression for the detection of transitional B cells (D), and for IgA and IgG expression for the detection of switched-memory B cells (E). P1 and P3 have fewer B cells than the healthy control analyzed in parallel (A). In addition, the IgMlow CD27−, the IgM+ CD27+ marginal-zone, and the IgM− CD27+ switched-memory B cell subsets are smaller, whereas the IgMhi CD27− B cell subset is larger, than in the control (B). BAFF-R expression is higher than in the control (BAFF-R mean fluorescence intensity of P1 = 2,476; P2 = 1,978; HD = 1,378). The proportion of CD10+ CD38+ transitional B cells in P1 is similar to that in the control (D), whereas IgA- or IgG-expressing switched-memory B cells are lacking (E).

Figure 4

Figure 4

Evaluation of Autophagy in LRBA-Deficient and Control Cells (A) In the top left panel, an image of a B cell from a control shows a normal ultrastructural morphology with well-defined mitochondria (M), Golgi apparatus (G), and ER (N, nucleus). An image from a B cell from P4 shows increased areas of Golgi apparatus (top right panel). Centrioles (arrow) were frequently found in the cytoplasm of B cells from P4 (bottom left panel), and accumulation of autophagosomes (indicated by white asterisks) in the cytoplasm was also frequently seen (bottom right panels). Scale bars represent 1 μm. (B) Autophagic flux showing the colocalization of LC3 and LysoID in the presence or absence of the lysosomal inhibitors E64d/pepstatin on PBMCs (Inh). Cells from P3 (red) show low levels of colocalization of LC3 and LysoID in all conditions tested. Error bars are shown and show the variation in LysoID and LC3 colocalization displayed by the B cells acquired on the Image Stream (n = 20,000). (C) Representative images of B cells from a healthy control and individual P3 show LC3 and lysoID colocalization.

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