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

. 2022 Sep 26;13:5644. doi: 10.1038/s41467-022-33285-x

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

© The Author(s) 2022

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. 3 — a HEK293FT cells were transfected with HA-IDO1, Flag-Ubiquitin, or MYC-USP14 as indicated. Polyubiquitination of IDO1 was then examined by immunoprecipitation with HA-beads and analyzed. Cells were treated with MG132 (10 µм) for 6 h before being harvested. b HCT116 cells treated with IU1 (50 µм) for 24 h or the cells stably transfected with control shRNA and USP14 shRNA were treated with MG132 (10 µм) for 6 h before being harvested. Lysates were immunoprecipitated with anti-IDO1 and analyzed. c HEK293FT cells were transfected with HA-K48-Ub and Flag-IDO1 with or without MYC-USP14 as indicated and cultured for 48 h. Cells were immunoprecipitated with anti-Flag and analyzed. d HCT116 cells were immunoprecipitated with anti-IDO1 or anti-TRIM21 and analyzed. e Flag-Ubiquitin was transfected together with HA-TRIM21 WT or HA-TRIM21 LD mutant in HEK293FT cells. Cells were treated with MG132 (10 µм) for 6 h before being harvested, then immunoprecipitated with anti-Flag and analyzed. f HCT116 cells with stable expression of control shRNA and TRIM21 shRNA were infected with Flag-IDO1. Cells were treated with MG132 (10 µм) for 6 h before being harvested, then immunoprecipitated with anti-Flag and analyzed. g Purified Flag-IDO1 was incubated with IgG control, purified HA-TRIM21 WT, or HA-TRIM21 LD mutant in the ubiquitination assay mix, and the sample was probed for the indicated proteins. h HA-Ub (WT, K48R, and K63R mutants) was transfected with or without Flag-TRIM21 in HEK293FT cells. Cells were immunoprecipitated with anti-HA and analyzed. i HCT116 cells were transfected with the indicated constructs and analyzed. j HEK293FT cells were transfected with IDO1, TRIM21, and Flag-tagged USP14 WT, C114A, or ΔUBL for 48 h and analyzed. k HA-TRIM21 was transfected with Flag-USP14 WT, C114A, or ΔUBL in HCT116 cells. Cells were treated with MG132 (10 µм) for 6 h before being harvested, then immunoprecipitated with anti-IDO1 and analyzed. The experiment was repeated three times independently with similar results (a–k). Source data are provided as a Source Data file.