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. 2021 Jul 12;10(7):655. doi: 10.3390/biology10070655

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© 2021 by the authors.

Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).

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Overexpression of MCPIP1 suppressed NF-κB activation in TNF-α-treated cells. T-REx-293 cells with (A) wild-type hemagglutinin (HA)-MCPIP1 or (B) the HA-MCPIP1-D141N mutant (HA-D141N) were pretreated without or with 1 μg/mL of doxycycline (Dox) for 16 h, and then treated without or with 10 ng/mL TNF-α for 30~90 min. Total cell lysates were collected and protein expressions were determined by Western blotting. (C,D) T-REx-293 cells with wild-type HA-MCPIP1 or the HA-MCPIP1-D141N mutant (HA-D141N) were pretreated without or with 1 μg/mL of Dox for 16 h, and then treated with 10 ng/mL TNF-α for (C) 15 or (D) 30 min. (C) Total cell lysates were collected, and IκB kinase (IKK) activity was determined by an in vitro kinase activity assay. (D) Cells were fixed and IF staining was performed with NF-κB p65 and HA antibodies, and DAPI was used as a counterstain for DNA.

Overexpression of MCPIP1 suppressed NF-κB activation in TNF-α-treated cells. T-REx-293 cells with (A) wild-type hemagglutinin (HA)-MCPIP1 or (B) the HA-MCPIP1-D141N mutant (HA-D141N) were pretreated without or with 1 μg/mL of doxycycline (Dox) for 16 h, and then treated without or with 10 ng/mL TNF-α for 30~90 min. Total cell lysates were collected and protein expressions were determined by Western blotting. (C,D) T-REx-293 cells with wild-type HA-MCPIP1 or the HA-MCPIP1-D141N mutant (HA-D141N) were pretreated without or with 1 μg/mL of Dox for 16 h, and then treated with 10 ng/mL TNF-α for (C) 15 or (D) 30 min. (C) Total cell lysates were collected, and IκB kinase (IKK) activity was determined by an in vitro kinase activity assay. (D) Cells were fixed and IF staining was performed with NF-κB p65 and HA antibodies, and DAPI was used as a counterstain for DNA.