Sample characterization.

<p>Sample characterization.</p

Correlations between CD25^bright/CD4⁺ aTreg frequencies and immunoblot band numbers.

<p>A–C: Bands recognized in immunoblots of HEp2-cytoplasmic proteins by SLE patients, unaffected relatives and unrelated control subjects. D–F: Bands recognized in immunoblots of HEp2-nuclear proteins. Regression lines represent linear regression.</p

Systematic gating for CD25^bright aTregs.

<p>A. First, in order to specify the upper limit of CD25 staining in conventional T-cells for each sample, the ninety-ninth percentile of PE/Cy5 fluorescence intensity within CD4⁻CD45RO⁻ cells (a population containing no Tregs) was determined as gate A. CD4⁺CD25^bright aTregs in the same sample were then quantified as those CD4⁺ cells that exceeded this value at least three-fold (gate B). B–D. The CD25^bright gate (B) contains high proportions of Foxp3+ cells in samples from SLE patients, unaffected relatives and healthy control subjects. As exemplified by an active SLE patient (panel B), an unaffected relative (panel C) and a healthy control subject (panel D), the CD25^bright gate B defined as 3xMFI(gate A) regularly contained 70–90% Foxp3+ cells within CD4+ lymphocytes when additional samples were stained for the same markers as previously but now in combination with intracellular Foxp3 staining.</p

Coreferentialities with CD25^bright/CD4⁺ frequencies.

*<p>Significance according to permutation test (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0033992#s4" target="_blank">methods</a>). Effects of bystander coreferentiality were checked for all significant tests (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0033992#s4" target="_blank">methods</a>), but rates of simulated bystander data reaching the respective test significance never exceeded 5%.</p

Group-wise coreferentialities between CD25^bright/CD4+ aTreg frequencies and <i>IL2RA</i> genetic-effects model scores.

<p>A: SLE patients, B: unaffected relatives, C: unrelated controls. Reactivities to cytoplasmic bands are indicated by filled circles, anti-nuclear reactivities by triangles and anti-brain reactivities by crosses.</p

Correlations between aTreg frequencies (CD25^bright/CD4⁺) and quantified SLE-associated specific autoreactive IgG.

<p>A–C: IgG anti-dsDNA. D–F: IgG anti-Sm. Regression lines represent linear regression.</p

Unweighted and family-weighted correlation coefficients.

<p>Distributions of unweighted and family-wighted correlation coefficients of CD25^bright/CD4⁺ aTreg frequencies (A) and log-transformed IgG anti-Sm (B) with all 130 single immunoblot reactivities are shown within SLE patients, unaffected relatives and unrelated controls, respectively. Reactivities to cytoplasmic bands are indicated by filled circles, anti-nuclear reactivities by triangles and anti-brain reactivities by crosses. Annoteted P-values were calculated by the described permutation test.</p

IgG autoreactivities in SLE patients, unafffected relatives and unrelated control subjects.

<p>A–F: Band numbers and 1st principal component calculated from HEp2 anti–cytoplasmic (A,B), anti-nuclear (C,D) and anti-brain (E,F) imunoblot reactivities. G–J: Specific SLE-associated autoreactive IgG quantified by ELISA. All plots show group-wise medians and results of pairwise Mann-Whitney tests for differences between groups.</p

Coreferentialities of the <i>IL2RA</i> model score with other parameters.

*<p>Test for multiple parameters, on maximal absolute coreferentiality |<i>R_C</i>|_max (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0033992#s4" target="_blank">methods</a>). Significant tests, except those for multiple SNPs, were also checked for the effect of bystander coreferentiality (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0033992#s4" target="_blank">methods</a>), but no such effect was detected.</p

Group-wise coreferentialities with aTreg frequencies, in respect to 130 IgG immunoblot reactivities as reference data.

<p>A–C: Coreferentiality between CD25^bright/CD4⁺ aTreg frequencies and IgG anti-HEp2-cytoplasmic immunoblot band numbers. D–F: Coreferentiality between aTreg frequencies and log-transformed IgG anti-Ro60/SSA. G–I: Coreferentiality between aTreg frequencies and log-transformed IgG anti-Sm. Reactivities to cytoplasmic bands are indicated by filled circles, anti-nuclear reactivities by triangles and anti-brain reactivities by crosses.</p