Figure 2 from Translational Activation of ATF4 through Mitochondrial Anaplerotic Metabolic Pathways Is Required for DLBCL Growth and Survival
Year: 2025
Type: other
Abstract: Knockdown SIRT3 caused ATF4 protein decrease via translation regulation. A, Western blots show ATF4 protein levels in different DLBCL cells with control or SIRT3 shRNAs. SIRT3 was blotted showing knockdown efficiency, and ACTB was used as reference protein control. B, qPCR results reflect the mRNA levels of ATF4 and PSAT1 in OCI-LY1 cells. Samples were harvested at day 4 after viral transduction. Results were normalized to the mRNA levels in control shRNA–transduced cells. C, qPCR results reflect the mRNA levels of ATF4 and PSAT1 in Karpas 422 cells. Samples were harvested at day 4 after viral transduction. Actin mRNA was used as reference, and results were normalized to the mRNA levels in control shRNA–transduced cells. D, qPCR results show the relative levels of ATF4 mRNAs in different cell lines at different time points after shRNA transduction. Samples were harvested at days 2, 3, 4, and 7 after viral transduction. Actin mRNA was used as reference, and results were normalized to the mRNA levels in control shRNA–transduced cells. E, Western blots show changes of phosphorylation of EIF2A and ATF4 protein levels in Karpas 422 cells with control or SIRT3 shRNAs. Total EIF2A and ACTB were blotted as loading controls. F, Western blots show GFP expression from the ATF4-5′UTR-GFP reporter and endogenous ATF4 protein levels in Karpas 422 cells with control or SIRT3 shRNAs. Tubulin and ACTB were blotted as loading controls. G, FCs of mean fluorescence intensity (MFI) of GFP expressed from the ATF4-5′UTR-GFP translation reporter in Karpas 422 cells with control or SIRT3 shRNAs. The data were collected from days 4 and 7 after viral transduction. MFI of normal yellow fluorescent protein (YFP) expression in control or SIRT3 knockdown cells were used for normalization to avoid background translation variations. H, FCs of MFI of GFP expressed from the ATF4-5′UTR-GFP translation reporter in OCI-LY1 cells with control or SIRT3 shRNAs. The data were collected from days 4 and 7 after viral transduction. MFI of normal YFP expression in control or SIRT3 knockdown cells were used for normalization to avoid background translation variations. Error bars represent the mean ± SD of three or more replicates.
Language: English
Subfield: Cell Biology
Domain: Life Sciences
Open Access status: gold
OpenAlex ID: W4416587420