STUB1 is essential for T-cell activation by ubiquitinating CARMA1
Article first published online: 15 FEB 2013
© 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
European Journal of Immunology
Volume 43, Issue 4, pages 1034–1041, April 2013
How to Cite
Wang, S., Li, Y., Hu, Y.-H., Song, R., Gao, Y., Liu, H.-Y., Shu, H.-B. and Liu, Y. (2013), STUB1 is essential for T-cell activation by ubiquitinating CARMA1. Eur. J. Immunol., 43: 1034–1041. doi: 10.1002/eji.201242554
- Issue published online: 16 APR 2013
- Article first published online: 15 FEB 2013
- Accepted manuscript online: 15 JAN 2013 07:53AM EST
- Manuscript Accepted: 11 JAN 2013
- Manuscript Revised: 3 DEC 2012
- Manuscript Received: 21 MAR 2012
- National Natural Science Foundation of China. Grant Numbers: 30700417, 30972719, 31170835, 30921001
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Figure S1. Knockdown of STUB1 inhibits NF-κB activation and IL-2 transcription upon anti-CD3/CD28 stimulation. (A) Jurkat E6 cells (5 × 106) stably transfected with control RNAi or STUB1 RNAi were challenged with anti-CD3/CD28 Abs as indicated. Cell lysates were analyzed by immunoblotting with the indicated Abs (A). The experiments were repeated for three times with similar results. RNA was isolated and mRNA levels of indicated genes were investigated by quantitative real-time PCR (B). The graph show means ± SD, n = 3 (* p < 0.05).
Figure S2. Knockdown of STUB1 inhibits the phosphorylation of IKK-α/β and TAK1 under P/I stimulation. Jurkat E6 cells (5 × 106) stably transfected with control RNAi or STUB1 RNAi were challenged with PMA/Ionomycin (P/I) (1 mMeach) as indicated. Cell lysates were analyzed by immunoblotting with the indicated Abs.
Figure S3. Interaction between overexpressed STUB1 and pathway members involved in TCR signaling. HEK293 cells (2 × 106) were transiently transfected with the indicated expression plasmids (5 μg each). Cell lysates were immunoprecipitated with anti-Flag and analyzed by immunoblotting with anti-HA mAb (upper). Expressions of the transfected proteins were analyzed by immunoblotting with anti-Flag and anti-HA mAbs (lower).
Figure S4. Knockdown of STUB1 has no marked effect on recruitment of BCL10 & MALT1 by CARMA1. Jurkat E6 cells (5 × 107) were challenged with P/I as indicated. Cell lysates were immunoprecipitated with anti-CARMA1. The immunoprecipitates were analyzed by immunoblotting with anti-CARMA1, anti-MALT1 and anti-BCL10 Abs. The expression levels of endogenous proteins were detected by immunoblotting with indicated antibodies respectively. The experiments were repeated for three times with similar results.
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