NBEAL2 is required for neutrophil and NK cell function and pathogen defense
John M. Sowerby, David C. Thomas, Simon Clare, Marion Espéli, Jose A. Guerrero, Kim Hoenderdos, Katherine Harcourt, Morgan Marsden, Juneid Abdul-Karim, Mathew Clement, Robin Antrobus, Yagnesh Umrania, Philippa R. Barton, Shaun M. Flint, Jatinder K. Juss, Alison M. Condliffe, Paul A. Lyons, Ian R. Humphreys, Edwin R. Chilvers, Willem H. Ouwehand, Gordon Dougan, Kenneth G.C. Smith
John M. Sowerby, David C. Thomas, Simon Clare, Marion Espéli, Jose A. Guerrero, Kim Hoenderdos, Katherine Harcourt, Morgan Marsden, Juneid Abdul-Karim, Mathew Clement, Robin Antrobus, Yagnesh Umrania, Philippa R. Barton, Shaun M. Flint, Jatinder K. Juss, Alison M. Condliffe, Paul A. Lyons, Ian R. Humphreys, Edwin R. Chilvers, Willem H. Ouwehand, Gordon Dougan, Kenneth G.C. Smith
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Brief Report Immunology Infectious disease

NBEAL2 is required for neutrophil and NK cell function and pathogen defense

Abstract

Mutations in the human NBEAL2 gene cause gray platelet syndrome (GPS), a bleeding diathesis characterized by a lack of α granules in platelets. The functions of the NBEAL2 protein have not been explored outside platelet biology, but there are reports of increased frequency of infection and abnormal neutrophil morphology in patients with GPS. We therefore investigated the role of NBEAL2 in immunity by analyzing the phenotype of Nbeal2-deficient mice. We found profound abnormalities in the Nbeal2-deficient immune system, particularly in the function of neutrophils and NK cells. Phenotyping of Nbeal2-deficient neutrophils showed a severe reduction in granule contents across all granule subsets. Despite this, Nbeal2-deficient neutrophils had an enhanced phagocyte respiratory burst relative to Nbeal2-expressing neutrophils. This respiratory burst was associated with increased expression of cytosolic components of the NADPH oxidase complex. Nbeal2-deficient NK cells were also dysfunctional and showed reduced degranulation. These abnormalities were associated with increased susceptibility to both bacterial (Staphylococcus aureus) and viral (murine CMV) infection in vivo. These results define an essential role for NBEAL2 in mammalian immunity.

Authors

John M. Sowerby, David C. Thomas, Simon Clare, Marion Espéli, Jose A. Guerrero, Kim Hoenderdos, Katherine Harcourt, Morgan Marsden, Juneid Abdul-Karim, Mathew Clement, Robin Antrobus, Yagnesh Umrania, Philippa R. Barton, Shaun M. Flint, Jatinder K. Juss, Alison M. Condliffe, Paul A. Lyons, Ian R. Humphreys, Edwin R. Chilvers, Willem H. Ouwehand, Gordon Dougan, Kenneth G.C. Smith

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Figure 3

Nbeal2–/– mice have dysfunctional NK cells in vitro and an impaired response to mCMV in vivo.

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Nbeal2–/– mice have dysfunctional NK cells in vitro and an impaired res...
(A) Proportion and absolute numbers of CD3–B220–CD49b+NKp46+ splenic NK cells for the indicated genotypes. (B) Expression of CD11b and CD27 (maturation markers) on CD3–B220–NK1.1+NKp46+ NK cells. Maturation was measured from gates R1 to R4 (with R4 being the most mature). Shown are the proportions and absolute numbers for each subset (n = 5–7). (C) Surface expression of LAMP-1/isotype of splenic NK cells 0 and 2 hours after PMA/ionomycin stimulation. Representative FACS gating and LAMP-1 histogram showing isotype control antibody staining (gray area) and LAMP-1 in Nbeal2+/+ (black line) and Nbeal2–/– mice (red line) (n = 4–5). max, maximum; MFI, mean fluorescence intensity. (D–F) Nbeal2+/+ or Nbeal2–/– mice were infected (i.p.) with 3 × 104 salivary gland–propagated Smith strain mCMV, and BW was monitored for 4 days after infection (D). On day 4, mice were sacrificed, and virus PFU were quantified in the spleen (E) and lungs (F) (n = 11). (G) Splenic NK cells were isolated and cultured in 1,000 U IL-2 for 4 days before cytotoxicity (LDH release) was tested on YAC-1 cells (n = 3). Data are presented as the mean and SD (A–D), median (E and F), or SEM (G) and are representative of 3 (A, E, and F) or 2 (B, C, and G) independent experiments. *P < 0.05, **P < 0.01, and ***P < 0.001, by Kruskal-Wallis test (A), Mann-Whitney U test (B, C, E, and F), or 2-way ANOVA (D and G).