A dominant-negative <i>IFNGR1</i> variant reveals broad immune cell sequestering of IFN-<b>γ</b>

Samantha Chan; Mai B. Margetts; Longfei Wang; Jack Godsell; Josh Chatelier; Belinda Liu; Charlotte A. Slade; Andrew Brett; Kasha P. Singh; Vanessa L. Bryant; Lauren J. Howson

doi:10.1172/JCI186799

A dominant-negative IFNGR1 variant reveals broad immune cell sequestering of IFN-γ

Samantha Chan,^1,2,3,4,5 Mai B. Margetts,¹ Longfei Wang,^2,6 Jack Godsell,^3,5 Josh Chatelier,^3,4 Belinda Liu,⁷ Charlotte A. Slade,^1,2,3 Andrew Brett,⁸ Kasha P. Singh,^9,10 Vanessa L. Bryant,^1,2,3 and Lauren J. Howson^1,2,3

Authorship note: Authors VLB and LJH are co–senior authors.

Published April 15, 2025 - More info

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To the Editor: IFN-γR1 deficiency is a form of Mendelian susceptibility to mycobacterial disease (MSMD) caused by partial or complete loss-of-function variants in IFNGR1 (1). Complete IFN-γR1 deficiency is autosomal recessive (AR) and characterized by complete penetrance, early onset, and severe infections (1, 2). Partial IFN-γR1 deficiency can be AR or autosomal dominant (AD) and typically has later onset, with less severe infections (1, 2). Dominant-negative IFN-γR1 deficiency is caused by variants in IFNGR1 exon 6 that result in a truncated receptor lacking both the intracellular internalization motif and the STAT1 docking site. This leads to surface accumulation of nonsignaling IFN-γ receptors that compete with WT receptors (1, 3). However, the implications of these nonsignaling receptors on the bioavailability of IFN-γ have not yet been explored. Here, we report an AD IFNGR1 variant in a family with MSMD and demonstrate the ubiquitous nature of IFN-γR1 expression and the capacity for dominant-negative IFN-γR1 variants to sequester IFN-γ on the cell surface.

Whole-exome sequencing confirmed a heterozygous IFNGR1 variant c.817delA (p.I273fs) present in family members with clinical disease (Figure 1A, see Supplemental Figure 1 and Supplemental Table 1 for patients’ clinical details; supplemental material available online with this article; https://doi.org/10.1172/JCI186799DS1). This variant occurs within exon 6 of IFNGR1, where previous dominant-negative variants have been reported (3).

Figure 1

IFN-γR1 is ubiquitously overexpressed and sequesters IFN-γ on the surface of patients’ cells. (A) Familial segregation of the IFNGR1 c.817delA (p.I273fs) variant. (B) Expression of IFN-γR1 on PBMC subsets. (C) pSTAT1 staining of PBMCs stimulated with IFN-γ and gated on monocytes. (D) PBMCs were cultured with LPS and a 10-fold dilution series of IFN-γ. (E) RNA-seq of differentially expressed immune genes. Surface IFN-γ detected on (F) monocytes and (G) PBMC subsets following incubation with IFN-γ. (H) IFN-γ dissociation from monocytes over time. The line represents nonlinear regression and the dashed line the dissociation half-life. Error bars represent SD between technical duplicates. HD, healthy donor; MAIT, mucosal-associated invariant T (cell). NS, no stimulation; P, patient.

IFN-γR1 is moderately expressed by almost every cell type in healthy individuals, and, in our patients, all PBMC subsets overexpressed IFN-γR1 (Figure 1B), having a 5–9-fold higher expression compared with individuals who were healthy (Supplemental Figure 2B). This is consistent with previously reported exon 6–truncated IFNGR1 variants (3) showing overexpression on monocytes and T cells, but furthers our understanding of the ubiquitous nature of patients’ IFN-γR1 overexpression to also encompass NK cells, B cells, and γδ T cells, and particularly high expression on MAIT cells.

It is suggested that IFN-γ signaling may be rescued by the addition of high-dose IFN-γ in dominant-negative IFN-γR1 deficiency (4). We observed a small dose-response effect to a maximum of one-fold increase in IFN-γR1^WT/I273fs monocyte pSTAT1 at 10 ng/mL IFN-γ that did not increase to healthy levels (7-fold) with increasing dose (Figure 1C). Addition of 0.1 ng/mL IFN-γ induced a maximum 1-fold increase in IFN-γR1^WT/I273fs monocyte LPS-induced TNF production, which did not increase to healthy levels (4-fold) with increasing dose (Figure 1D). RNA-seq analysis confirmed that IFN-γR1^WT/I273fs monocyte sensitivity to IFN-γ could not be rescued with high-dose exposure across downstream gene targets of IFN-γ signaling (Figure 1E).

We next investigated IFN-γR1 binding kinetics by culturing PBMCs with IFN-γ and measuring IFN-γR1 and IFN-γ by flow cytometry. Upon binding IFN-γ, the WT receptor decreased 2.5-fold at the surface and 1.4-fold intracellularly (Supplemental Figure 2C). The IFN-γR1^WT/I273fs cells, with an overall higher level of IFN-γR1 baseline expression, showed impaired surface (0.7-fold decrease) and intracellular (0.5-fold decrease) decreases, suggesting an impaired degradation due to absence of the internalization domain in these variant receptors. As expected, IFN-γ was undetectable on IFN-γR1^WT/WT monocytes, due to the WT receptor internalization following IFN-γ binding (Figure 1F). However, IFN-γR1^WT/I273fs monocytes showed a dose-dependent increase in surface IFN-γ at concentrations up to 1,000 ng/mL (Figure 1F). IFN-γ was highest on monocytes, but all PBMC subsets exhibited detectable dose-dependent increases (Figure 1G). We then measured IFN-γ cytokine-receptor dissociation by preincubating patient PBMCs with IFN-γ and measuring surface IFN-γ on monocytes over time. We observed prolonged detection of IFN-γ on the cell surface, with a dissociation half-life of 2 hours (Figure 1H).

Broad sequestering of IFN-γ on the patients’ cell surface has the potential to reduce systemic IFN-γ bioavailability in affected patients. This may include endogenous IFN-γ, providing an explanation for the low plasma IFN-γ in patients with AD IFN-γR1 deficiency, while IFN-γ plasma levels in AR IFN-γR1 are typically either moderate (partial deficiency) or high (complete deficiency) (5, 6). It may also impact the bioavailability of exogenous IFN-γ, such as the recombinant therapy used to treat acute refractory mycobacterial infection. IFN-γ treatment has been reported to be effective in certain cases of dominant-negative IFN-γR1 anecdotally (summarized in Supplemental Table 2); however, no studies have directly assessed the efficacy of IFN-γ therapy for patients with dominant-negative IFN-γR1 deficiency.

In summary, we have demonstrated impaired IFN-γ signaling in AD IFN-γR1 deficiency that cannot be rescued by high-dose IFN-γ in vitro. This is potentially due to prolonged surface retention of IFN-γ by ubiquitously overexpressed truncated IFN-γR1, which establishes an inaccessible reservoir of IFN-γ sequestered on cell surfaces in dominant-negative IFN-γR1 deficiency.

References

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[1] van de Vosse E, van Dissel JT. IFN-γR1 defects: Mutation update and description of the IFNGR1 variation database. Hum Mutat. 2017;38(10):1286–1296.
View this article via: CrossRef PubMed Google Scholar

[2] Dorman SE, et al. Clinical features of dominant and recessive interferon gamma receptor 1 deficiencies. Lancet. 2004;364(9451):2113–2121.
View this article via: CrossRef PubMed Google Scholar

[3] Jouanguy E, et al. A human IFNGR1 small deletion hotspot associated with dominant susceptibility to mycobacterial infection. Nat Genet. 1999;21(4):370–378.
View this article via: CrossRef PubMed Google Scholar

[4] Holland SM. Treatment of infections in the patient with Mendelian susceptibility to mycobacterial infection. Microbes Infect. 2000;2(13):1579–1590.
View this article via: PubMed Google Scholar

[5] Fieschi C, et al. High levels of interferon gamma in the plasma of children with complete interferon gamma receptor deficiency. Pediatrics. 2001;107(4):E48.
View this article via: CrossRef PubMed Google Scholar

[6] Sologuren I, et al. Partial recessive IFN-γR1 deficiency: genetic, immunological and clinical features of 14 patients from 11 kindreds. Hum Mol Genet. 2011;20(8):1509–1523.
View this article via: CrossRef PubMed Google Scholar

A dominant-negative IFNGR1 variant reveals broad immune cell sequestering of IFN-γ

Samantha Chan,^1,2,3,4,5 Mai B. Margetts,¹ Longfei Wang,^2,6 Jack Godsell,^3,5 Josh Chatelier,^3,4 Belinda Liu,⁷ Charlotte A. Slade,^1,2,3 Andrew Brett,⁸ Kasha P. Singh,^9,10 Vanessa L. Bryant,^1,2,3 and Lauren J. Howson^1,2,3

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A dominant-negative IFNGR1 variant reveals broad immune cell sequestering of IFN-γ

Samantha Chan,1,2,3,4,5 Mai B. Margetts,1 Longfei Wang,2,6 Jack Godsell,3,5 Josh Chatelier,3,4 Belinda Liu,7 Charlotte A. Slade,1,2,3 Andrew Brett,8 Kasha P. Singh,9,10 Vanessa L. Bryant,1,2,3 and Lauren J. Howson1,2,3

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Samantha Chan,^1,2,3,4,5 Mai B. Margetts,¹ Longfei Wang,^2,6 Jack Godsell,^3,5 Josh Chatelier,^3,4 Belinda Liu,⁷ Charlotte A. Slade,^1,2,3 Andrew Brett,⁸ Kasha P. Singh,^9,10 Vanessa L. Bryant,^1,2,3 and Lauren J. Howson^1,2,3