Figure 6.
Capping-independent stimulation of the LARP7–7SK binding by MePCE, which leads to the stabilization of 7SK by the cooperative actions of MePCE and LARP7. (A) Gel mobility shift assay was performed with 32P-labeled wild-type 7SK or 7SK (Δ4U’s) and in the presence (+) or absence (−) of the indicated proteins or SAM. The positions of free 7SK as well as the various 7SK-containing complexes are indicated on the left. (B) 7SK capping reactions containing the same concentration (1×) as or twice (2×) or thrice (3×) more MePCE than that used in gel shift reactions in A were performed in the presence of SAM and absence of LARP7. The RNA products were isolated and analyzed by autoradiography (top panel) and EB staining (bottom panel). (C) HeLa cells were transfected with constructs expressing the indicated shRNAs to KD the expression of MePCE or/and LARP7. The levels of 7SK and the other indicated proteins in NEs of transfected cells were analyzed by WB and NB.