Figure - PMC

Skip to main content

An official website of the United States government

Here's how you know

Here's how you know

Official websites use .gov
A .gov website belongs to an official government organization in the United States.

Secure .gov websites use HTTPS
A lock ( ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

View full-text article in PMC

. 2018 Oct 30;9:4515. doi: 10.1038/s41467-018-06924-5

Search in PMC
Search in PubMed
View in NLM Catalog
Add to search

© The Author(s) 2018

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

PMC Copyright notice

Fig. 2 — SUMOylation of ROR-γt inhibits IL-17 expression. a Data show heatmap visualization of upregulated genes determined by RNA seq analysis in ROR-γt^–/– CD4⁺ T cells transduced with either WT ROR-γt or K187R-ROR-γt and differentiated under Th17-inducing conditions. b Real-time PCR analysis was conducted for IL-17a mRNA expression in ROR-γt^–/– CD4⁺ T cells transduced with either WT ROR-γt or K187R-ROR-γt. The relative fold change in the mRNA levels of the genes was normalized against ROR-γt^–/– cells as compared with lentivirus-transduced ROR-γt^–/– cells. c ELISA was conducted for IL-17A secretion in the culture supernatant of ROR-γt^–/– cells transduced with lentivirus encoding V5-tagged WT-ROR-γt or K187R-ROR-γt. d Immunoblot analysis of the expression of lentiviral constructs of V5-ROR-γt in ROR-γt^–/– cells. e Luciferase assay was conducted of lysates from Jurkat T cells transfected with various combinations (below plot) of the IL-17-promoter-driven luciferase plasmid (pGL4-mIL17p) and plasmid-encoded HA-SUMO1, Myc-Ubc9, Flag-ROR-γt, and/or Flag-K187R-ROR-γt. f Luciferase assay was conducted of lysates from Jurkat T cells transfected with various combinations (below plot) of plasmid pGL4-mIL17p along with Flag-ROR-γt, HA-SUMO1, and either Myc-Ubc9 or Myc-Ubc9-C93A. Results are presented in relative luciferase units (RLU). g Data show Ubc9 knockdown by Ubc9-specific shRNA (shUbc9) or control shRNA (shCtrl) in WT cLPLs that were stimulated with PMA and ionomycin. ELISA was performed to measure IL-17A secretion in culture supernatant of Ubc9 knockdown cLPLs as well as immunoblotting showing knockdown of Ubc9. h ELISA was performed to measure the IL-17A secretion in the culture supernatant of Ubc9 knockdown cells expressing either WT-ROR-γt or K187R-ROR-γt. i Real-time PCR analysis for IL-17a mRNA expression, in which the relative fold change in the mRNA levels of the genes has been normalized against ROR-γt-expressing cells treated with control shRNA compared to Ubc9-specific shRNA. j Immunoblot analysis shows Ubc9 knockdown in CD4⁺ T cells expressing either WT-ROR-γt or K187R-ROR-γt. Data are from one experiment representative of three independent experiments with similar results. *p < 0.05, **p < 0.01, ns: non-significant (two-tail t test) error bars are S.D.