The effect of miR-30 on target mRNA adenylation status after manipulation of active PABP. The LM-PAT assay was used to test poly(A) tail length (see Materials and Methods for details). 293T cells were transfected under conditions described in Figure 4, A or D. (A) The effects of PABP and PABP (M161A) ectopic overexpression on the poly(A) tail length of the miR-30 8× target mRNA 20 h post-transfection. (B) miR-30 8× target mRNA assayed as in A at 12 h post-transfection. (C) The adenylation status of a control mRNA (GAPDH) was assayed with the highest amount (800 ng) of PABP and PABP (M161A) cDNA transfected at 20 h and 12 h post-transfection. (D) LM-PAT assays of miR-30 8× target mRNA (upper panel) and control mRNA (GAPDH; lower panel) after knockdown of Paip2.
