Fig. 3. Effects of cAMP on actin (A), PKA RII (B), fusion (C) and acidification (D) in J774 macrophages. (A) Effects of indicated treatments on LBP-associated actin. Micrograph shows the effect of actin on intracellular LBPs visualized by Rhodamine-phalloidin labeling with a magnification of the LBP area in untreated cells. (Left) DIC-image; (right) Rhodamine-phalloidin image. The histogram shows percentage of LBPs positive for actin under indicated conditions. (B) Analysis of the co-localization of PKA RII. Micrograph shows DIC image of LBPs in macrophages (left), immunofluorescent labeling of PKA RII on LBPs (middle) and the overlay (right). The histogram shows the percentage of LBPs co-localizing with PKA RII under indicated conditions. The blot shows identification of PKA RII on LBPs. (C) Analysis of fusion of LBPs with Rhodamine-gold-filled late endosomes and lysosomes. Micrograph shows an example of lysosomal Rhodamine-gold particles co-localizing with LBPs with a magnification of the LBP area in untreated cells: DIC image (left), Rhodamine-gold image (right). The histogram shows the percentage of LBPs co-localizing with lysosomal Rhodamine-gold under indicated conditions. (D) Analysis of phagosomal acidification. The histogram shows the percentage of LBPs that co-localize with LysoTracker Red DND-99 under indicated conditions. All histograms represent mean ± s.d. of three individual experiments. Asterisks indicate significance as determined by the Student's t-test: ^*P<0.01 compared with levels in the control.