PMC6302768_01
(A) Neonatal pustulosis. (B) Cutis laxa. (C) Timeline for the clinical manifestations and treatments of the patient. IRT, Immunoglobulin replacement therapy.
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F1
She presented with very severe vesiculo-pustular rash in the 1st week of life (Figure 1A). Cutis laxa was evident since she was 6 years-old (Figure 1B) and progressive sensorineural deafness was diagnosed at the age of 7, requiring a hearing-assistive device. Supplementary Table 1 ). She started prophylactic co-trimoxazole, immunoglobulin replacement, as well as low dose steroids (5 mg every other day) and hydroxychloroquine (Figure 1C), which led to substantial improvement.
Reduced production of IL-10 and IL-1beta after LPS stimulation; reduced production of IFN-gamma and IL-17 in response to polyclonal T-cell stimulation (performed at 9, 10, and 11 years-old).
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F2
Whole blood stimulation assays showed a strongly reduced production of IFN-gamma and IL-17 in response to polyclonal T-cell stimulation and reduced production of IL-10 and IL-1beta after LPS stimulation (Figure 2).
(A,B)
De novo L848P mutation in the PLCG2 gene; (C) Leu848 is conserved in most vertebrates; (D) L848P mutation is predicted to affect the PLCgamma2 structure while L848F had only minimal impact; (E) Both L848P (3) and S707Y (4) mutants showed increased basal and EGF-stimulated activity in comparison to the wild-type PLCgamma2 (2) and control COS cells (1).
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F3
Sanger sequencing showed that this mutation appeared de novo in the patient, while being absent from both parents and a healthy sibling (Figures 3A,B). The mutation affected amino acid residue Leu848 in the split pleckstrin homology (spPH) domain of the PLCgamma2 protein that is conserved in most vertebrates (Figure 3C). We used I-Tasser to model the PLCgamma2 protein fragment between amino acids 770 and 1044 containing the L848P and L848F substitutions and found that L848P is predicted to affect the PLCgamma2 structure, while L848F had only minimal impact (Figure 3D). IP production. In this standard cellular assay, both L848P and S707Y mutants showed increased basal and EGF-stimulated activity in comparison to wild-type PLCgamma2 (Figure 3E).