Fig. 2: RG4 inhibits TMPRSS2 translation.
From: RNA G-quadruplex in TMPRSS2 reduces SARS-CoV-2 infection
a Levels of endogenous TMPRSS2 protein (left panel), and mRNA (right panel) in H1299 cells treated with PDS (10 μM), cPDS (10 μM), or TMPyP4 (10 μM). ImageJ quantification of the TMPRSS2/ACTIN ratio is shown (bottom panel). b The full-length ORF region of human Tmprss2 or its RG4-mutant sequence was inserted into the MCS of pcDNA3.1 vector. The bold letters represent the G-tracts and the underlined letters represent mutations. c Levels of HA-TMPRSS2 protein (left panel), and mRNA (right panel) in H1299 cells transfected with hG4WT or hG4mut1 plasmids. ImageJ quantification of the HA-TMPRSS2/ACTIN ratio is shown (bottom panel). d, e Levels of HA-TMPRSS2 protein (left panel), and mRNA (right panel) in H1299 cells transfected with hG4WT (d) or hG4mut1 (e) plasmids, and stimulated with PDS (10 μM), cPDS (10 μM), or TMPyP4 (10 μM). ImageJ quantification of the HA-TMPRSS2/ACTIN ratio is shown (bottom panel). f Levels of HA-TMPRSS2 protein (left panel), and mRNA (right panel) in hACE2-293T cells transfected with hG4WT or hG4mut1 plasmids. ImageJ quantification of the HA-TMPRSS2/ACTIN ratio is shown (bottom panel). g, h Levels of HA-TMPRSS2 protein (left panel), and mRNA (right panel) in hACE2-293T cells transfected with hG4WT (g) or hG4mut1 (h) plasmids, and stimulated with PDS (10 μM), cPDS (10 μM), or TMPyP4 (10 μM). ImageJ quantification of the HA-TMPRSS2/ACTIN ratio is shown (bottom panel). i Polysome shift analysis of endogenous Tmprss2 mRNA in H1299 cells. Representative polysome gradient profile shows the position of polysomes, monosomes (80 S), small (40 S), and large (60 S) subunits (left panel). Tmprss2 expression in control and PDS-treated cells was calculated as the percentage of mRNA in each fraction compared with total mRNA of all fractions (right panel). j, k Polysome shift analysis of exogenous Tmprss2 mRNA in hG4WT (j) and hG4mut1 (k) transfected hACE2-293T cells. A representative polysome gradient profile was shown on the left. Tmprss2 expression in each fraction from control and PDS-treated cells was shown on the right. Data are shown as mean ± SEM, n = 3. Two-tailed Student’s t test. Source data are provided as a source data file.
