Inherited GATA2 Deficiency Is Dominant by Haploinsufficiency and Displays Incomplete Clinical Penetrance

Abstract

Purpose: Germline heterozygous mutations of GATA2 underlie a variety of hematological and clinical phenotypes. The genetic, immunological, and clinical features of GATA2-deficient patients with mycobacterial diseases in the familial context remain largely unknown.

Methods: We enrolled 15 GATA2 index cases referred for mycobacterial disease. We describe their genetic and clinical features including their relatives.

Results: We identified 12 heterozygous GATA2 mutations, two of which had not been reported. Eight of these mutations were loss-of-function, and four were hypomorphic. None was dominant-negative in vitro, and the GATA2 locus was found to be subject to purifying selection, strongly suggesting a mechanism of haploinsufficiency. Three relatives of index cases had mycobacterial disease and were also heterozygous, resulting in 18 patients in total. Mycobacterial infection was the first clinical manifestation in 11 patients, at a mean age of 22.5 years (range: 12 to 42 years). Most patients also suffered from other infections, monocytopenia, or myelodysplasia. Strikingly, the clinical penetrance was incomplete (32.9% by age 40 years), as 16 heterozygous relatives aged between 6 and 78 years, including 4 older than 60 years, were completely asymptomatic.

Conclusion: Clinical penetrance for mycobacterial disease was found to be similar to other GATA2 deficiency-related manifestations. These observations suggest that other mechanisms contribute to the phenotypic expression of GATA2 deficiency. A diagnosis of autosomal dominant GATA2 deficiency should be considered in patients with mycobacterial infections and/or other GATA2 deficiency-related phenotypes at any age in life. Moreover, all direct relatives should be genotyped at the GATA2 locus.

Keywords: GATA2; Primary immunodeficiency; haploinsufficiency; mycobacteria; tuberculosis.

Figure 1.. GATA2 deficiency.

Schematic representation of the 15 kindreds with germline GATA2 mutations. Each kindred is designated by a capital letter (A-O), each generation by a Roman numeral (I-II), and each individual by an Arabic numeral. The proband is indicated by arrows. Solid black shapes indicate GATA2-deficient patients and clinical disease. Solid gray shapes indicate individuals with GATA2 deficiency-related clinical features (lymphedema or acute myeloid leukemia [AML]) who have not been genotyped. Mutations are indicated in red and wild type (WT) allele in black. Individuals whose genetic status could not be determined are denoted “E?” Individuals with symbols crossed by a black vertical line have asymptomatic GATA2 deficiency. Deceased individuals are represented by symbols crossed by a diagonal black line. Healthy individuals are shown in white.

Figure 2.. Protein expression of GATA2 alleles in an overexpression system.

HEK293T cells were transfected with an empty plasmid (EV), a construct encoding GATA2 wild-type (WT)-DDK-tagged or mutated alleles corresponding to isoform 1 of GATA2 (p.R330*, p.M388T, p.R396W, p.R396Q, p.R398Q, p.R398W, c.599dup, c.915_916del, c.1023del, c.1035_1036insTCTGGCC, c.1099dup, p.V382Gfs*23, p.N381_V382ins41 and p. S340_N381del) for 48 hours. GATA2 protein levels were evaluated by western blotting with antibodies against GATA2 and against the DDK tag. GAPDH was used as a loading control. NT indicates non-transfected cells. The results shown are representative of three independent assays.

Figure 3.. Functional characterization of GATA2 alleles.

A. GATA2-luciferase activity in HEK293T cells. GATA2-dependent transactivation potential was evaluated with a GATA2-luciferase reporter plasmid. Cells were transiently transfected with (+) or without (–) PML, empty plasmid (EV) and various GATA2-plasmids (WT, p.R330*, p.M388T, p.R396W, p.R396Q, p.R398Q, p.R398W, c.599dup, c.915_916del, c.1023del, c.1035_1036insTCTGGCC, c.1099dup, p.V382Gfs*23, p.N381_V382ins41 and p. S340_N381del). B. Transient cotransfection of cells with PML and equivalent amounts of GATA2-WT plasmid (25 ng) but with various amounts of mutant plasmids (p.R330*, p.M388T, p.R396W, p.R396Q, p.R398Q, p.R398W, c.599dup, c.915_916del, c.1023del, c.1035_1036insTCTGGCC, c.1099dup, p.V382Gfs*23, p.N381_V382ins41 and p. S340_N381del), with different amounts of EV used to keep the final plasmid concentration constant. Error bars indicate the standard error of the mean. The results shown are representative of two independent assays with three replicates per experiment.

Figure 4.. List of variants and strength of the purifying selection acting on GATA2.

Genome-wide distribution of the strength of purifying selection, estimated by A. Probability of loss of function intolerance (pLI) score, B. the f parameter, C. residual variation intolerance (RVIS) score on bar plots, as described in the methods and D. consensus negative selection (CoNeS) score. The position of GATA2 is indicated by a black arrow and a red vertical bar.

Figure 5.. Mycobacterial infections and distribution in GATA2-deficient patients.

A. Spectrum of mycobacterial species causing disease in GATA2-deficient patients. The patients infected with a particular “type” of mycobacterial species are represented as percentages, with infections with two different species considered as a type of infection. Asymptomatic carriers are individuals carrying the mutation with no clinical phenotype. B. Mycobacterial infection sites in patients infected with EM infection.

Figure 6.. Penetrance of clinical diseases in patients with GATA2 deficiency.

A. Kaplan-Meier curves showing the penetrance of GATA2 in confirmed carriers (GATA2 confirmed, continuous line) and family members with suspected GATA2 deficiency (GATA2 suspected, discontinuous line) with clinical manifestations at 20 and 40 years. B. Penetrance of mycobacterial infection (mycobacterial disease, continuous line) and other clinical manifestations of GATA2 deficiency (other disease, discontinuous line). C. Kaplan-Meier curve showing the percentage of mycobacterial infection-free patients from 5 to 45 years. The total number of patients (n=16) is indicated by a continuous black line, patients infected with environmental mycobacteria (EM, n=11) are indicated by a discontinuous gray line, patients infected with Mycobacterium tuberculosis (TB, n=4) are indicated by a discontinuous black line and patients infected with unidentified mycobacteria (Mycobacterium spp., n=3) are indicated in a black dotted line.