Huang, L., W. Xu, H. Liu, M. Xue, X. Liu, K. Zhang, L. Hu, J. Li, X. Liu, Z. Xiang, J. Zheng, C. Li, W. Chen, Z. Bu, T. Xiong, and C. Weng. 2021. African swine fever virus pI215L negatively regulates cGAS-STING signaling pathway through recruiting RNF138 to inhibit K63-linked ubiquitination of TBK1. J. Immunol. 207: 2754–2769.

Several of the panel labels were switched in the legend for Fig. 3 as originally published. The part of the legend originally labeled as (D)–(F) referred to panels (A)–(C), and the part of the legend labeled as (A)–(C) referred to panels (D)–(F). The figure was correct as published. The corrected legend for Fig. 3 is shown below.

FIGURE 3. ASFV pI215L inhibits K63-linked polyubiquitination and phosphorylation of TBK1. (A–C) HEK293T cells were transfected with an IFN-β luciferase (Luc) reporter and a Renilla-TK reporter, a plasmid expressing HA-STING (A), HA-TBK1 (B), or HA-IRF3-5D (C), together with an empty vector or different amounts (0, 100, 200, and 400 ng) of a plasmid expressing Flag-pI215L for 24 h, then cells were collected for detection of the Luc activity. The expression of HA-STING, HA-TBK1, HA-IRF3-5D, Flag-pI215L, and GAPDH was detected by Western blotting (WB). (D–F) HEK293T cells were transfected with a plasmid expressing HA-STING (D), HA-TBK1 (E), or HA-IRF3-5D (F), together with an empty vector or increasing amounts of a plasmid expressing Flag-pI215L. The cells were collected 24 h posttransfection and the mRNA levels of IFN-β were analyzed by qPCR. The expression of HA-STING, HA-TBK1, HA-IRF3-5D, Flag-pI215L, and GAPDH were detected by Western blotting. (G) HEK293T cells were transfected with a plasmid expressing Flag-pI215L or an empty vector, and then the cells were infected with HSV-1 for 12 h. The cell lysates were then analyzed with the indicated Abs. (H) Quantitation of TBK1-p/TBK1 ratio from the ImageJ analysis in (G). n = 3; error bars, SD. (I–L) Co-IP analysis of the polyubiquitination of TBK1 (I) and IRF3 (J), and the K63-linked (K) and K48-linked (L) polyubiquitination of TBK1. HEK293T cells were transfected with plasmids expressing Flag-TBK1, Flag-IRF3, Myc-ubiquitin (Ub), or Myc-Ub-K63 or Myc-Ub-K48 with or without HA-pI215L. The cell lysates were immunoprecipitated (IP) with anti-Flag (M2) beads for detection of TBK1 ubiquitination. The proteins were detected with indicated Abs by Western blotting. Data are representative of three independent experiments with three biological replicates. ***p < 0.001 (one-way ANOVA).

In addition, a revised version of Fig. 8 correcting the 293T-WT labels to be 293T-Rnf138+/+ in panels (I) and (J) was not incorporated into the published version of the article. The corrected version of Fig. 8 is shown below. The figure legend was correct as published and is shown below for reference.

FIGURE 8.
FIGURE 8. ASFV pI215L promotes RNF138 to degrade RNF128. (A) HEK293T cells were transfected with a plasmid encoding Flag-RNF128 alone or together with different doses of a plasmid encoding HA-pI215L. The expression of pI215L and RNF128 was analyzed by Western blotting (WB). (B) Quantitation of RNF128/b-tubulin ratio from ImageJ analysis in (A). n 5 3; error bars, SD. (C) HEK293T cells were transfected with a plasmid encoding HARNF128, along with different doses of a plasmid encoding Flag-RNF138. The expression of RNF138 and RNF128 was analyzed by Western blotting. (D) Quantitation of RNF128/b-tubulin ratio from ImageJ analysis in (C). n 5 3; error bars, SD. (E) HEK293T cells were transfected with a plasmid encoding HA-RNF128 alone or together with a plasmid encoding Flag-RNF138 and then treated with or without MG132. The expression of pI215L and RNF128 was analyzed by Western blotting. (F) Quantitation of RNF128/b-tubulin ratio from ImageJ analysis in (E). n 5 3; error bars, SD. (G) HEK293T cells were transfected with a plasmid encoding Flag-RNF128 alone or together with a plasmid encoding Flag-pI215L or Flag-RNF138 or both. The expression of pI215L, RNF138, and RNF128 was analyzed by Western blotting. (H) Quantitation of RNF128/b-actin ratio from ImageJ analysis in (J). n 5 3; error bars, SD. (I) HEK293T or HEK293T-Rnf138−/− cells were transfected with a plasmid expressing Flag-pI215L or HA-RNF128 alone or both together. The expression of pI215L and RNF128 was analyzed by Western blotting. (J) Quantitation of RNF128/GAPDH ratio from ImageJ analysis in (I). n 5 3; error bars, SD. (K) HEK293T or HEK293T-Rnf138−/− cells were transfected with an empty vector or a plasmid expressing Flag-pI215L. The expression of pI215L, RNF138, and RNF128 was analyzed by Western blotting. (L) Quantitation of RNF128/GAPDH ratio from ImageJ analysis in (K). n 5 3; error bars, SD. (M) PAMs were transfected with scramble siRNA or siRNA targeting Rnf138 and then mock infected or infected with ASFV for 24 h. The expression of endogenous pI215L, RNF138, and RNF128 was analyzed by Western blotting. (N) Quantitation of RNF128/GAPDH ratio from ImageJ analysis in (M). n 5 3; error bars, SD. 0.01 &lt; *p &lt; 0.05, 0.001 &lt; **p &lt; 0.01, ***p &lt; 0.001 (one-way ANOVA).
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ASFV pI215L promotes RNF138 to degrade RNF128. (A) HEK293T cells were transfected with a plasmid encoding Flag-RNF128 alone or together with different doses of a plasmid encoding HA-pI215L. The expression of pI215L and RNF128 was analyzed by Western blotting (WB). (B) Quantitation of RNF128/b-tubulin ratio from ImageJ analysis in (A). n 5 3; error bars, SD. (C) HEK293T cells were transfected with a plasmid encoding HARNF128, along with different doses of a plasmid encoding Flag-RNF138. The expression of RNF138 and RNF128 was analyzed by Western blotting. (D) Quantitation of RNF128/b-tubulin ratio from ImageJ analysis in (C). n 5 3; error bars, SD. (E) HEK293T cells were transfected with a plasmid encoding HA-RNF128 alone or together with a plasmid encoding Flag-RNF138 and then treated with or without MG132. The expression of pI215L and RNF128 was analyzed by Western blotting. (F) Quantitation of RNF128/b-tubulin ratio from ImageJ analysis in (E). n 5 3; error bars, SD. (G) HEK293T cells were transfected with a plasmid encoding Flag-RNF128 alone or together with a plasmid encoding Flag-pI215L or Flag-RNF138 or both. The expression of pI215L, RNF138, and RNF128 was analyzed by Western blotting. (H) Quantitation of RNF128/b-actin ratio from ImageJ analysis in (J). n 5 3; error bars, SD. (I) HEK293T or HEK293T-Rnf138−/− cells were transfected with a plasmid expressing Flag-pI215L or HA-RNF128 alone or both together. The expression of pI215L and RNF128 was analyzed by Western blotting. (J) Quantitation of RNF128/GAPDH ratio from ImageJ analysis in (I). n 5 3; error bars, SD. (K) HEK293T or HEK293T-Rnf138−/− cells were transfected with an empty vector or a plasmid expressing Flag-pI215L. The expression of pI215L, RNF138, and RNF128 was analyzed by Western blotting. (L) Quantitation of RNF128/GAPDH ratio from ImageJ analysis in (K). n 5 3; error bars, SD. (M) PAMs were transfected with scramble siRNA or siRNA targeting Rnf138 and then mock infected or infected with ASFV for 24 h. The expression of endogenous pI215L, RNF138, and RNF128 was analyzed by Western blotting. (N) Quantitation of RNF128/GAPDH ratio from ImageJ analysis in (M). n 5 3; error bars, SD. 0.01 < *p < 0.05, 0.001 < **p < 0.01, ***p < 0.001 (one-way ANOVA).

FIGURE 8.
FIGURE 8. ASFV pI215L promotes RNF138 to degrade RNF128. (A) HEK293T cells were transfected with a plasmid encoding Flag-RNF128 alone or together with different doses of a plasmid encoding HA-pI215L. The expression of pI215L and RNF128 was analyzed by Western blotting (WB). (B) Quantitation of RNF128/b-tubulin ratio from ImageJ analysis in (A). n 5 3; error bars, SD. (C) HEK293T cells were transfected with a plasmid encoding HARNF128, along with different doses of a plasmid encoding Flag-RNF138. The expression of RNF138 and RNF128 was analyzed by Western blotting. (D) Quantitation of RNF128/b-tubulin ratio from ImageJ analysis in (C). n 5 3; error bars, SD. (E) HEK293T cells were transfected with a plasmid encoding HA-RNF128 alone or together with a plasmid encoding Flag-RNF138 and then treated with or without MG132. The expression of pI215L and RNF128 was analyzed by Western blotting. (F) Quantitation of RNF128/b-tubulin ratio from ImageJ analysis in (E). n 5 3; error bars, SD. (G) HEK293T cells were transfected with a plasmid encoding Flag-RNF128 alone or together with a plasmid encoding Flag-pI215L or Flag-RNF138 or both. The expression of pI215L, RNF138, and RNF128 was analyzed by Western blotting. (H) Quantitation of RNF128/b-actin ratio from ImageJ analysis in (J). n 5 3; error bars, SD. (I) HEK293T or HEK293T-Rnf138−/− cells were transfected with a plasmid expressing Flag-pI215L or HA-RNF128 alone or both together. The expression of pI215L and RNF128 was analyzed by Western blotting. (J) Quantitation of RNF128/GAPDH ratio from ImageJ analysis in (I). n 5 3; error bars, SD. (K) HEK293T or HEK293T-Rnf138−/− cells were transfected with an empty vector or a plasmid expressing Flag-pI215L. The expression of pI215L, RNF138, and RNF128 was analyzed by Western blotting. (L) Quantitation of RNF128/GAPDH ratio from ImageJ analysis in (K). n 5 3; error bars, SD. (M) PAMs were transfected with scramble siRNA or siRNA targeting Rnf138 and then mock infected or infected with ASFV for 24 h. The expression of endogenous pI215L, RNF138, and RNF128 was analyzed by Western blotting. (N) Quantitation of RNF128/GAPDH ratio from ImageJ analysis in (M). n 5 3; error bars, SD. 0.01 &lt; *p &lt; 0.05, 0.001 &lt; **p &lt; 0.01, ***p &lt; 0.001 (one-way ANOVA).
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ASFV pI215L promotes RNF138 to degrade RNF128. (A) HEK293T cells were transfected with a plasmid encoding Flag-RNF128 alone or together with different doses of a plasmid encoding HA-pI215L. The expression of pI215L and RNF128 was analyzed by Western blotting (WB). (B) Quantitation of RNF128/b-tubulin ratio from ImageJ analysis in (A). n 5 3; error bars, SD. (C) HEK293T cells were transfected with a plasmid encoding HARNF128, along with different doses of a plasmid encoding Flag-RNF138. The expression of RNF138 and RNF128 was analyzed by Western blotting. (D) Quantitation of RNF128/b-tubulin ratio from ImageJ analysis in (C). n 5 3; error bars, SD. (E) HEK293T cells were transfected with a plasmid encoding HA-RNF128 alone or together with a plasmid encoding Flag-RNF138 and then treated with or without MG132. The expression of pI215L and RNF128 was analyzed by Western blotting. (F) Quantitation of RNF128/b-tubulin ratio from ImageJ analysis in (E). n 5 3; error bars, SD. (G) HEK293T cells were transfected with a plasmid encoding Flag-RNF128 alone or together with a plasmid encoding Flag-pI215L or Flag-RNF138 or both. The expression of pI215L, RNF138, and RNF128 was analyzed by Western blotting. (H) Quantitation of RNF128/b-actin ratio from ImageJ analysis in (J). n 5 3; error bars, SD. (I) HEK293T or HEK293T-Rnf138−/− cells were transfected with a plasmid expressing Flag-pI215L or HA-RNF128 alone or both together. The expression of pI215L and RNF128 was analyzed by Western blotting. (J) Quantitation of RNF128/GAPDH ratio from ImageJ analysis in (I). n 5 3; error bars, SD. (K) HEK293T or HEK293T-Rnf138−/− cells were transfected with an empty vector or a plasmid expressing Flag-pI215L. The expression of pI215L, RNF138, and RNF128 was analyzed by Western blotting. (L) Quantitation of RNF128/GAPDH ratio from ImageJ analysis in (K). n 5 3; error bars, SD. (M) PAMs were transfected with scramble siRNA or siRNA targeting Rnf138 and then mock infected or infected with ASFV for 24 h. The expression of endogenous pI215L, RNF138, and RNF128 was analyzed by Western blotting. (N) Quantitation of RNF128/GAPDH ratio from ImageJ analysis in (M). n 5 3; error bars, SD. 0.01 < *p < 0.05, 0.001 < **p < 0.01, ***p < 0.001 (one-way ANOVA).

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In the second sentence under the Cell viability assays heading in the Materials and Methods, “stimulated with cGAMP” and “infected with or without ASFV” were switched. The corrected Cell viability assays paragraph is shown below.

Cell viability was assessed by the MTT assay (30). Briefly, PAMs cultured in 96-well plates were infected with or without ASFV at a multiplicity of infection (MOI) of 1 and then mock stimulated or stimulated with cGAMP for 4 h, 8 h, 12 h, and 24 h, respectively, and then 200 µl DMEM containing 20 µl MTT (5 mg/ml) was added to each well and incubated for 4 h at 37°C. The culture medium was then removed, and 100 µl DMSO was added to each well, and the plate was agitated on an orbital shaker for 15 min. The light absorbance in each sample was measured at 570 nm with an ELISA reader.

The items above have been corrected in the online version of the article, which now differs from the print version as originally published.

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Copyright © 2022 by The American Association of Immunologists, Inc.
2022
Copyright © 2022 by The American Association of Immunologists, Inc.