Fig 1: Marginal zone (MZ) B cells express Gpr43 and Gpr43-/- mice have heightened levels of serum immunoglobulin M (IgM) specific to double-stranded DNA (dsDNA) and phosphatidylcholine (PC). (a) Messenger RNA from fluorescence-activated cell sorting-purified follicular B (Fo B) or MZ B cells from Gpr43 +/+ and Gpr43 -/- mice was subjected to quantitative reverse transcription-PCR analysis to measure the expression of Gpr40, Gpr41, Gpr43, Gpr120 andGpr109a. All data are shown as mean values ± s.e.m. of two independent experiments (n = 4 mice per group), except for Gpr109a which was quantified in a separate experiment with n = 4 Gpr43 +/+ and n = 6 Gpr43 -/-. (b) Splenocytes of Gpr43 +/+ and Gpr43 -/- mice were analyzed by flow cytometry. B220+CD93+ immature or B220+CD93- mature B-cell population are gated. B-cell precursors within the immature population were further divided into T1 (IgM+CD23-) or T2 (IgM+CD23+) cells. The mature B-cell population was further stained for CD21 and CD23 to divide cells into Fo B (CD21-CD23+) or MZ B cells (CD21+CD23-). B-1 cells were identified as CD19+B220-/low cells. Bar graphs show the frequencies or absolute cell numbers of T1, T2, Fo B, MZ B and B-1 cells. (c) Splenocytes from Gpr43 +/+ and Gpr43 -/- mice were stained for MZ B cell markers and mean fluorescence intensity (MFI) levels of the indicated surface molecules are plotted. Data are shown as mean values ± s.e.m. of two independent experiments (n = 5 or 6 mice per group). (d) Immunofluorescence staining of spleen sections from 10- to 12-week-old Gpr43 +/+ and Gpr43 -/- mice with B220 (blue) for B cells, CD1d (red) for MZ B cells and MOMA-1 (green) for marginal metallophilic macrophages. The MZ B-cell layer is marked by white arrows. Slides are representative of four mice per group. (e, f) The serum of 15 Gpr43 +/+ and 12 Gpr43 -/- 10–15-week-old naïve mice was screened for the presence of total IgM, IgG, IgG3 and IgG1 at various serum dilutions (e) and dsDNA- and PC-specific IgG and IgM at serum concentration of 1:50 (f). The Mann–Whitney U-test was used for comparison, *P < 0.05, **P < 0.01, ****P < 0.0001.