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[Preprint]. 2024 May 25:2024.05.24.595774. [Version 1] doi: 10.1101/2024.05.24.595774

Figure 4. ALTO downregulates MCPyV early gene expression via NF-κB signaling.

Figure 4.

A) DNA pulldowns with MKL-2 HA-ALTO cell lysates incubated with biotinylated MCPyV NCCR or AmpR dsDNA coupled to streptavidin beads. B) MCPyV early luciferase in 293A cells transfected with HA-ALTO wt, HA-ALTO ΔNTAR1+2 or HA-RelA plasmids. Luciferase activity was normalized to CMV-driven renilla activity. Statistical significance was calculated by one-way ANOVA. * P<0.03, ** P<0.002. C) Western blots from 293A cells transfected with either HA-ALTO wt; HA-ALTO ΔNTAR1+2; HA-RelA; HA-RelB and FLAG-p52; or HA-RelA, HA-RelB and FLAG p52 plasmids. D) qRT-PCR analysis of relative LT and ST expression normalized to 36B4 expression in MKL-2 RFP, HA-ALTO wt or ΔNTAR1+2 cells treated with doxycycline for 6 days. The data show the mean ± the standard deviation of three technical replicates. Statistical significance was calculated by 2-way ANOVA. * P<0.03, ** P<0.003, *** P<0.0002, **** P<0.0001. E) Western blots of MKL-2 cells expressing RFP, HA-ALTO wt or ΔNTAR1+2 harvested after 6 days doxycycline induction. F) Growth curves of MKL-2 cells expressing RFP, HA-ALTO wt or ΔNTAR1+2. Each data point represents the mean ± standard deviation of three or more technical replicates. Statistical significance was calculated by 2-way ANOVA. **** P<0.0001.