|
|
Blood, Vol. 109, Issue 10, 4368-4375, May 15, 2007
Nonredundant roles for Stat5a/b in directly regulating Foxp3
Blood Yao et al.
109: 4368
Supplemental materials for: Yao et al, Vol 109, Issue 10, 4368-4375
Files in this Data Supplement:
- Figure S1. Specificity of Foxp3 staining in CD4+ T cells (PDF, 16.5 KB) -
Flow cytometric analysis using anti- Foxp3 antibody versus an isotype control (IgG2a) in wild-type thymic CD4+ SP cells. The cells were fixed, permeablized, and stained with Foxp3 antibody or isotype control as indicated.
- Figure S2. Reduction but not absence of Stat5a/b in T cells from Stat5fl/–, CD4cre, Yfp mice (PDF, 25.8 KB) -
(A) YFP expression in thymic and splenic CD4+ SP cells from Stat5fl/–, CD4cre, Yfp (heavy line) and Stat5fl/fl (light line) mice. Stat5a/b mRNA (B) and protein expression (C) were assessed in sorted CD4 SP T cells from Stat5fl/fl mice and Stat5fl/–, CD4cre, Yfp mice. For Stat5 protein expression, the images were juxtaposed, removing additional controls.
- Figure S3. Autoimmune disease in Stat5fl/–, CD4cre, Yfp mice (PDF, 55 KB) -
Organs were removed from 5- to 7-week-old Stat5fl/–, CD4cre, Yfp mice. Tissues were stained with hematoxylin and eosin. Mononuclear cell infiltrates are noted in heart, liver, lung, and stomach.
- Figure S4. Role of Stat3 in negatively regulating Foxp3 (PDF, 24.8 KB) -
(A) Stat3 mRNA levels were assessed in splenic CD4+ T cells from Stat3fl/fl and Stat3fl/fl, MMTVcre mice. (B) Sorted splenic CD25–CD4+ T cells from Stat3fl/fl and Stat3fl/fl, MMTVcre mice were cultured with plate-bound anti-CD3 and anti-CD28 in combination with TGF -1 or TGF-1 plus IL-6. Regulation of Foxp3+ by IL-6 in CD4+ T cells from Stat3fl/fl versus Stat3fl/fl, MMTVcre mice; Means ± SE (n = 3) are shown; *P < .01. (C) Stat3 marginally binds the Foxp3 gene. Sorted CD25+CD4+ splenic T cells were treated with IL-6 for 1 hour. Proteins and DNA were cross-linked with formaldehyde, cells were lysed, and DNA was sheared. Chromatin immunoprecipitation was performed using either normal rabbit serum or anti-Stat3 antibody. Quantification of immunoprecipitated DNA fragments by real-time PCR using primers and probes for sites I, II, III, and IV (as defined in Figure 7) was performed.
| |
