FIG. 9.
PMA-activated MIE gene expression is partially dependent on cellular NF-κB subunits p65 and p50. NT2 cells were transfected with a plasmid expressing shRNA against p65 (p65iY and p65iB), p50 (p50iY and p50iB), RelB (RelBiY, RelBiG, and RelBiB), or a negative control (NCi) for 72 h. These cells were then infected for 24 h and stimulated with PMA by using the approach described in the legend of Fig. 4A. The expression levels of MIE proteins (IE1 p72 and IE2 p86) were analyzed by Western blotting at 22 h poststimulation. Tubulin served as an internal control. Relative to NCi, p65iY and p65iB decreased levels of p65 by 80% and 78%, respectively, and decreased IE1 p72 levels by 56% and 47%, respectively. p50iY and p50iB decreased p50 levels by 63% and 77%, respectively, and decreased IE1 p72 levels by 77% and 95%, respectively; p50iB also decreased the p65 level by 61%, whereas p50iY decreased the p65 level by 31%. All results are normalized to tubulin levels.