Loss of ARHGEF1 causes a human primary antibody deficiency
Amine Bouafia, … , Eric Oksenhendler, Sven Kracker
Amine Bouafia, … , Eric Oksenhendler, Sven Kracker
Published December 6, 2018
Citation Information: J Clin Invest. 2019;129(3):1047-1060. https://doi.org/10.1172/JCI120572.
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Research Article Immunology

Loss of ARHGEF1 causes a human primary antibody deficiency

Abstract

ARHGEF1 is a RhoA-specific guanine nucleotide exchange factor expressed in hematopoietic cells. We used whole-exome sequencing to identify compound heterozygous mutations in ARHGEF1, resulting in the loss of ARHGEF1 protein expression in 2 primary antibody–deficient siblings presenting with recurrent severe respiratory tract infections and bronchiectasis. Both ARHGEF1-deficient patients showed an abnormal B cell immunophenotype, with a deficiency in marginal zone and memory B cells and an increased frequency of transitional B cells. Furthermore, the patients’ blood contained immature myeloid cells. Analysis of a mediastinal lymph node from one patient highlighted the small size of the germinal centers and an abnormally high plasma cell content. On the molecular level, T and B lymphocytes from both patients displayed low RhoA activity and low steady-state actin polymerization (even after stimulation of lysophospholipid receptors). As a consequence of disturbed regulation of the RhoA downstream target Rho-associated kinase I/II (ROCK), the patients’ lymphocytes failed to efficiently restrain AKT phosphorylation. Enforced ARHGEF1 expression or drug-induced activation of RhoA in the patients’ cells corrected the impaired actin polymerization and AKT regulation. Our results indicate that ARHGEF1 activity in human lymphocytes is involved in controlling actin cytoskeleton dynamics, restraining PI3K/AKT signaling, and confining B lymphocytes and myelocytes within their dedicated functional environment.

Authors

Amine Bouafia, Sébastien Lofek, Julie Bruneau, Loïc Chentout, Hicham Lamrini, Amélie Trinquand, Marie-Céline Deau, Lucie Heurtier, Véronique Meignin, Capucine Picard, Elizabeth Macintyre, Olivier Alibeu, Marc Bras, Thierry Jo Molina, Marina Cavazzana, Isabelle André-Schmutz, Anne Durandy, Alain Fischer, Eric Oksenhendler, Sven Kracker

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Figure 3

Compound heterozygous mutations in ARHGEF1 lead to protein deficiency in the patients’ lymphocytes.

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Compound heterozygous mutations in ARHGEF1 lead to protein deficiency in...
(A) Pedigree of the 2 siblings presenting with PAD. (B) Sanger sequencing of ARHGEF1 in P1, P2, and their parents. A nonsense mutation c.898C>T (Chr19: 42398710: C>T; hg19 build 137) was inherited from the father, and a splice mutation c.1669-1G>T (Chr19: 42406933: G>T) was inherited from the mother. F, father; M, mother. (C) Impact of ARHGEF1 mutations on the protein sequence. (D) RT-PCR analysis of the presence of ARHGEF1 transcripts lacking exon 19 (Δ exon 19) in the patients’ PBMCs, their mother, and a healthy donor (HD). L, ladder; H2O, water control. The schema for PCR analyses is depicted here; arrows designate primers used to analyze the effect of the c.1669-1G>T mutation on ARHGEF1 exon 19 splicing. The absence of the transcript lacking exon 19 was verified in 2 blood samples from independent healthy donors. (E and F) Western blots showing the expression of ARHGEF1 and RhoA in protein lysates of (E) B-EBV and (F) T cell blasts derived from patients. GAPDH was included as a loading control. (G and H) Enzyme-linked immunosorbent assay of the level of active RhoA (RhoA-GTP) in (G) B-EBV cells and (H) T cell blasts derived from patients. The level of active RhoA in the patients’ cells was compared with that found in HD-derived cells. In G, each symbol indicates an independent measure. Two independent healthy donor–derived B-EBV cell lines (HD1, circles, n = 3; HD2, white squares, n = 1), 2 independent B-EBV cell lines from P1 (P1-1, black squares, n = 3; P1-2, crosses, n = 2), and 1 B-EBV cell line from P2 (P2, triangles, n = 3) were analyzed. *P < 0.05; **P < 0.01, 1-sample, 2-tailed t test on normalized log2-transformed measurements. The experiment presented in H was performed only once.