KHSRP promotes miR-155 maturation, but inhibits miR-155* production. (A) Anti-KHSRP antibody immunoprecipitates pri-miR-155. Primary human PDCs were treated with R837 for indicated times and lysed, and total cell extracts were immunoprecipitated, as indicated. Control immunoglobulin G (IgG; Ctrl-IgG) was used as the negative control. RNA was purified from immunocomplexes and analyzed by qRT-PCR. (B-D) Human PDCs transfected with either the control siRNA or KHSRP siRNA were stimulated with R837 over a 16-hour time course and harvested at the indicated time points. miR-155 (B) and miR-155* (C) were analyzed with the TaqMan MicroRNA Expression Assay and normalized to the RNU48 levels. Pri-miR-155 mRNA (D) was detected by qRT-PCR and normalized to the RPL13a RNA levels. These graphs show the fold induction calculated by normalizing the expression values at different time points to the control 0-hour values. (E) Immunoblot kinetics analysis for KHSRP. An α-tubulin immunoblot is shown for equal loading control. Data are representative of 2 independent experiments. (F) KHSRP was induced by R837 and IFN-α. Kinetic analysis of the induction of KHSRP mRNA by R837 and IFN-α. Human PDCs were stimulated with IFN-α or R837 alone or in the presence of 5 μg/mL type I IFN blocking receptor B18R (anti-IFN) over a 24-hour time course and harvested at the indicated time points. KHSRP mRNA was detected by qRT-PCR and normalized to RPL13a RNA levels. The graph shows the fold induction calculated by normalizing the expression values at different time points to the 0-hour values. The data are representative of at least 3 independent experiments, each based on a different PDC preparation.