Figure 7.

DNASE1L3-sensitive Ags on MP include DNA-associated proteins. (A) Human plasma IgG binding to proteins in MPs from Jurkat T cells. Plasma samples from healthy controls (Ctrl), DNASE1L3-deficient patients (1L3 null), sporadic SLE patients (SLE), or purified IgG from a healthy control (Ctrl IgG) and sporadic SLE patient (SLE IgG) were tested by Western blot. The presence (+) or absence (−) of IgG binding to DNASE1L3-sensitive MP Ags as assessed by flow cytometry is indicated. (B) Histones and HMGB1 as potential Ags on MPs as determined by Western blot. Plasma samples from patients with sporadic SLE were analyzed for IgG binding to MP proteins as in A. In parallel, MPs were probed by anti-H2A, anti-H2B, and anti-HMGB1 Abs. Representative of two independent experiments. (C and D) SLE-prone mouse plasma IgG binding to MP proteins as determined by Western blot. In C, plasma from a Dnase1l3-deficient mouse (1L3 KO mouse, dilutions 1:50, 1:150, and 1:450) and plasma from a patient with sporadic SLE (SLE human, dilutions 1:150, 1:500, and 1:1,500) were tested in parallel. In D, plasma from Dnase1l3-deficient (1L3 KO) mice before (3 mo) or after the onset of SLE-like disease (7–10 mo) was tested. Representative of two independent experiments. (E) DNASE1L3-sensitive exposure of HMGB1 and DNA on the surface of Jurkat MPs. MPs that were untreated or pretreated with recombinant human DNASE1 or DNASE1L3 were stained for HMGB1 (top) or DNA (Vybrant Green, bottom) and analyzed by flow cytometry. Representative of three independent experiments. SSC-A, side scatter signal area.