Mutations in GATA2 cause human NK cell deficiency with specific loss of the CD56(bright) subset

Abstract

Mutations in the transcription factor GATA2 underlie the syndrome of monocytopenia and B- and natural killer (NK)-cell lymphopenia associated with opportunistic infections and cancers. In addition, patients have recurrent and severe viral infections. NK cells play a critical role in mediating antiviral immunity. Human NK cells are thought to mature in a linear fashion, with the CD56(bright) stage preceding terminal maturation to the CD56(dim) stage, considered the most enabled for cytotoxicity. Here we report an NK cell functional defect in GATA2-deficient patients and extend this genetic lesion to what is considered to be the original NK cell-deficient patient. In most cases, GATA2 deficiency is accompanied by a severe reduction in peripheral blood NK cells and marked functional impairment. The NK cells detected in peripheral blood of some GATA2-deficient patients are exclusively of the CD56(dim) subset, which is recapitulated on in vitro NK cell differentiation. In vivo, interferon α treatment increased NK cell number and partially restored function but did not correct the paucity of CD56(bright) cells. Thus, GATA2 is required for the maturation of human NK cells and the maintenance of the CD56(bright) pool in the periphery. Defects in GATA2 are a novel cause of profound NK cell dysfunction.

Figures

Figure 1

GATA2-deficient patients have defective NK cell cytotoxicity. NK cell functional activity was measured against susceptible K562 target cells in the presence of IL-2 where indicated (dashed line) using PBMCs isolated from whole blood. PBMCs from 5 patients (blue) were evaluated in combination with gender-matched controls (red).

Figure 2

Reduced frequency of CD56+CD3− NK cells and enrichment of the CD56dim NK cell subset in GATA2 patients. Analysis of PBMCs from whole blood by FACS. Cells were (A) gated on lymphocytes, (B) further gated on CD56+CD3−, and (C) then analyzed for CD56 density with the aid of CD16 to identify CD56dim populations. (Bottom) One representative healthy donor control is shown.

Figure 3

Expression of NK cell surface markers. Analysis of NK cells from whole blood using the gating strategy described in Figure 2. Shown are percent positive of CD56+CD3− NK cells for each GATA2-deficient patient and corresponding healthy donor controls with each point representing the value from a single subject. The large horizontal bar denotes the mean and the vertical bar demonstrates the standard deviation. Excluded from analysis are those patients with <1% of NK cells in peripheral blood.

Figure 4

Expression of GATA2 in mature NK cells. B cells, T cells, monocytes, and NK cells (all 106) or 5 × 105 CD56bright or CD56dim NK cells were isolated from peripheral blood of a healthy donor by FACS, lysed, and immunoblotted for (A) GATA2 and (B) actin as a loading control. Shown is 1 of 3 representative experiments. B, CD19+ B cells; M, CD14+ monocytes; MW, molecular weight protein ladder; NK, CD56+CD3− NK cells; T, CD3+ T cells.

Figure 5

GATA2 is required for NK cell differentiation from CD34+ precursors. Highly purified CD34+ hematopoietic precursors from GATA2-deficient patient 3 or a healthy donor were isolated from peripheral blood by FACS and cultured on EL08.1D2 stromal cells in the presence of IL-3, IL-6, IL-7, stem cell factor, Flt3L, and IL-15. After 30 days, cells were isolated and (A) expansion was calculated based on cell number recovered for control (black) and patient (white). (B) Patient cells (black) and control cells (red) were analyzed for CD56 expression. Control cells were 2.3% CD56bright. Isotype control staining is shown in gray. (C) CD56 on (left) control cells or (right) patient cells was analyzed in combination with pan-KIR antibody (bottom).

Figure 6

In vivo–administered IFNα increases NK cell numbers and functionality but does not restore CD56bright insufficiency. PBMCs were isolated from whole blood of patients 7 and 8 before and after initiation of IFNα2b treatment. (A) Patient (blue) or control (red) NK cell cytotoxicity was assayed against susceptible K562 target cells (left) before and (right) after treatment in the presence (dashed) or absence of IL-2 (solid). (B) PBMCs from whole blood were isolated and analyzed by FACS as in Figure 2 (top) before and (bottom) after treatment.