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

. 2020 Jun 19;11(12):894–914. doi: 10.1007/s13238-020-00734-6

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

© The Author(s) 2020

Open AccessThis 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 licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence 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 licence, visit http://creativecommons.org/licenses/by/4.0/.

PMC Copyright notice

TRIM35 catalyzes K48-linked polyubiquitination and degradation of IAV PB2. (A) Co-IP and IB analysis of HEK293T cells expressing TRIM35-V5 and WSN (H1N1) PB2. 10 μg/mL MG132 was added into cells at 6 h post-transfection (right panel). (B) Co-IP and IB analysis of HEK293T cells expressing TRAF3-Flag, TRIM35-V5, with or without WSN (H1N1) PB2. (C) IB analysis of HEK293T cells expressing WSN (H1N1) PB2 and increasing amounts of TRIM35-V5. (D) IB analysis of HEK293T cells expressing TRIM35-V5 or control vector for 36 h, followed by infection with WSN (H1N1) virus. (E) Co-IP and IB analysis to assess WSN (H1N1) PB2 ubiquitination in HEK293T cells expressing WSN (H1N1) PB2, with or without TRIM35-V5, along with HA-ubiquitin (K48) or HA-ubiquitin (K48R). (F) Co-IP and IB analysis to assess WSN (H1N1) PB2 ubiquitination in HEK293T cells expressing WSN (H1N1) PB2, with or without TRIM35-V5. (G) Minigenome assay in HEK293T cells expressing WSN (H1N1) PB2, PB1, PA, and NP, pHH21-SC09NS F-Luc, pRL-TK, along with control vector, TRIM35-V5, or both TRIM35-V5 and HA-ubiquitin. Results are expressed relative to Renilla luciferase activity. (H) Co-IP and IB analysis to assess WSN (H1N1) PB2 ubiquitination in HEK293T cells expressing WSN (H1N1) PB2, HA-ubiquitin (K48), along with V5-tagged TRIM35 or TRIM35 DelRING mutant. (I) IB analysis of HEK293T cells expressing V5-tagged TRIM35 DelRING mutant or control vector for 36 h, followed by infection with WSN (H1N1) virus. (J) Replication of WSN (H1N1) virus (MOI = 0.1) in HEK293 cells transfected for 24 h to express V5-tagged TRIM35 DelRING mutant or control vector, determined by plaque assay on MDCK cells. IB analysis was shown in the bottom panel. (K) Co-IP and IB analysis to assess K48-linked ubiquitination of WSN (H1N1) PB2 or its mutants in HEK293T cells expressing V5-tagged WSN (H1N1) PB2 or its mutants, GST-TRIM35, and HA-ubiquitin (K48). (L) IB analysis of HEK293T cells expressing WSN (H1N1) PB2 K736R mutant and increasing amounts of TRIM35-V5. Data are representative of at least three independent experiments. Means ± SD are shown in (G) and (J, upper panel) (n = 3). Two-tailed unpaired t-test was used for the statistical analysis, ***P < 0.001