Journal: iScience
Article Title: SQSTM1/p62 Controls mtDNA Expression and Participates in Mitochondrial Energetic Adaption via MRPL12
doi: 10.1016/j.isci.2020.101428
Figure Lengend Snippet: SQSTM1/p62 induces MRPL12 Expression via Activating p38/ATF2 Signaling Pathway (A) Schematic diagram for SQSTM1/p62 domains and interacting partners. ① to ⑥ represent inhibitors or activators to the interacting partners as following: ① p38 inhibitor SB203580 (10 μM, Selleck); ② Nrf2 activator NK252 (20 μM, MedChemExpress); ③mTOR inhibitor rapamycin (10 μM, Selleck); ④ IRS-1/2 inhibitor NK157 (10 μM, Selleck); ⑤ ERK1/2 activator Honokiol (20 μM, Selleck); and ⑥ PKC inhibitor Go6983 (100 nM, Selleck). (B) MRPL12 expression was detected by western blotting in SQSTM1/p62-overexpressed HK-2 cells treated with the above inhibitors or activators. (C) A potential ATF2 binding site within the promoter region of human MRPL12 gene was identified by ALGGEN-PROMO (Version 3.0.2) between −2,344 bp and −2,336 bp upstream from the transcription start site. (D–G) Western blotting for total p38 and ATF2 in SQSTM1/p62-overexpressed (D) or knocked down (F) HK-2 cells. GAPDH was used as internal control. (E) and (G) were grayscale analysis for (D) and (F) respectively. Unpaired t tests were used, n = 3, ∗∗p < 0.01. (H–K) Western blotting for nuclear p-p38 and p-ATF2 in SQSTM1/p62-overexpressed (H) or knocked down (J) HK-2 cells. LaminB was used as internal control. (I) and (K) were grayscale analysis for (H) and (J) respectively. Unpaired t tests were used, n = 3, ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001. (L and M) Immunofluorescent staining for SQSTM1/p62 and p-ATF2 in SQSTM1/p62 transfected HK-2 cells. Nuclei were counterstained by Hoechst 33258. Scale bars, 20 μm. Mean intensity was calculated by Image Pro Plus and showed in histogram respectively. Unpaired t tests were used, n = 3, ∗p < 0.05, ∗∗p < 0.01.
Article Snippet: 1 to 6 represent inhibitors or activators to the interacting partners as following: 1 p38 inhibitor SB203580 (10 μM, Selleck); 2 Nrf2 activator NK252 (20 μM, MedChemExpress); 3mTOR inhibitor rapamycin (10 μM, Selleck); 4 IRS-1/2 inhibitor NK157 (10 μM, Selleck); 5 ERK1/2 activator Honokiol (20 μM, Selleck); and 6 PKC inhibitor Go6983 (100 nM, Selleck). (B) MRPL12 expression was detected by western blotting in SQSTM1/p62-overexpressed HK-2 cells treated with the above inhibitors or activators. (C) A potential ATF2 binding site within the promoter region of human MRPL12 gene was identified by ALGGEN-PROMO (Version 3.0.2) between −2,344 bp and −2,336 bp upstream from the transcription start site. (D–G) Western blotting for total p38 and ATF2 in SQSTM1/p62-overexpressed (D) or knocked down (F) HK-2 cells.
Techniques: Expressing, Western Blot, Binding Assay, Control, Staining, Transfection
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