HCMV infection downregulates GPX4 and stimulates lipid peroxidation but does not induce ferroptosis Figure 2

FIG 2 GPX4 regulation occurs in part through the 3′UTR. (A) Western blot analysis of lysates from uninfected MRC-5 cells and MRC-5 cells infected with TB40/E-WT virus at MOI = 3, treated with vehicle control (−) or acyclovir (+), and harvested at 24 or 96 hpi. Blots were probed for GPX4, IE1, IE2, pp28, and p115 as a loading control. The values under the GPX4 blot denote quantification of each GPX4 band in the blot, relative to the GPX4 band in the uninfected cells. (B) Western blot analysis of lysates from uninfected MRC-5 cells or MRC-5 cells infected with TB40/E-WT virus or UV-inactivated TB40/E-WT virus at MOI = 3 and harvested at 4, 8, 12, 24, and 120 hpi. Blots were probed for GPX4, IE1, IE2, and p115 as a loading control. The values under the GPX4 blot denote quantification of each GPX4 band in the blot, relative to the GPX4 band in the uninfected cells. (C) qPCR analysis of GPX4 transcripts isolated from uninfected or TB40/E-WT infected human dermal fibroblasts (MOI = 3) at 24, 48, 72, 96, and 120 hpi. GPX4 transcripts were normalized to GAPDH and shown as fold change relative to the uninfected sample. Results are from four independent experiments. (D) Figure portraying the cloning of a control SV40 terminator sequence or GPX4 3′UTR sequence into the luciferase plasmid. The GPX4 3′UTR sequence is shown with notable elements and miRNA-binding sites marked. SECIS denotes the selenocysteine insertion sequence, a component of the GPX4 sequence that is required for its translation. (E) Luciferase activity of MRC-5 cells expressing a luciferase control plasmid or a luciferase GPX4 3′UTR plasmid and either uninfected or infected with TB40/E-WT at MOI = 3 and harvested at 24 hpi. Luciferase activity is shown relative to uninfected MRC-5 cells expressing the luciferase control plasmid. Results are from two independent experiments, n = 6. For the above panels, * indicates P < 0.05 and ns = not significant.

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Norbury, Christopher (2024). HCMV infection downregulates GPX4 and stimulates lipid peroxidation but does not induce ferroptosis Figure 2 [Data set]. Scholarsphere.

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Work Title HCMV infection downregulates GPX4 and stimulates lipid peroxidation but does not induce ferroptosis Figure 2
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  1. Christopher Norbury
License CC BY 4.0 (Attribution)
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Publication Date December 20, 2024
Deposited December 20, 2024

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    • FIG 2 GPX4 regulation occurs in part through the 3′UTR. (A) Western blot analysis of lysates from uninfected MRC-5 cells and MRC-5 cells infected with TB40/E-WT virus at MOI = 3, treated with vehicle control (−) or acyclovir (+), and harvested at 24 or 96 hpi. Blots were probed for GPX4, IE1, IE2, pp28, and p115 as a loading control. The values under the GPX4 blot denote quantification of each GPX4 band in the blot, relative to the GPX4 band in the uninfected cells. (B) Western blot analysis of lysates from uninfected MRC-5 cells or MRC-5 cells infected with TB40/E-WT virus or UV-inactivated TB40/E-WT virus at MOI = 3 and harvested at 4, 8, 12, 24, and 120 hpi. Blots were probed for GPX4, IE1, IE2, and p115 as a loading control. The values under the GPX4 blot denote quantification of each GPX4 band in the blot, relative to the GPX4 band in the uninfected cells. (C) qPCR analysis of GPX4 transcripts isolated from uninfected or TB40/E-WT infected human dermal fibroblasts (MOI = 3) at 24, 48, 72, 96, and 120 hpi. GPX4 transcripts were normalized to GAPDH and shown as fold change relative to the uninfected sample. Results are from four independent experiments. (D) Figure portraying the cloning of a control SV40 terminator sequence or GPX4 3′UTR sequence into the luciferase plasmid. The GPX4 3′UTR sequence is shown with notable elements and miRNA-binding sites marked. SECIS denotes the selenocysteine insertion sequence, a component of the GPX4 sequence that is required for its translation. (E) Luciferase activity of MRC-5 cells expressing a luciferase control plasmid or a luciferase GPX4 3′UTR plasmid and either uninfected or infected with TB40/E-WT at MOI = 3 and harvested at 24 hpi. Luciferase activity is shown relative to uninfected MRC-5 cells expressing the luciferase control plasmid. Results are from two independent experiments, n = 6. For the above panels, * indicates P < 0.05 and ns = not significant.
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    • 2024-12-20
  • Added Creator Christopher Norbury
  • Added Maddie Martin 2022-11-09 09h53m23s.scn
  • Added Maddie Martin 2022-11-10 11h29m19s.scn
  • Added Maddie Martin 2022-11-09 10h00m08s.scn
  • Added Maddie Martin 2022-11-10 11h34m24s.scn
  • Added protein calcs GPX4 acyclovir & UV.xlsx
  • Added 2022-02-15 GPX4 FSP1 timecourses.xlsx
  • Added 2022-02-18 GPX4 FSP1 timecourses 2.xls
  • Added EF1a Luc combined.xlsx
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    • https://creativecommons.org/licenses/by/4.0/
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