(A) Heat map of all significantly down regulated targets in a microarray study using LNCaP cells transiently expressing miR-331-3p. All targets are represented with or without a predicted miR-331-3p seed region. (B) Further stratification of the most down regulated targets identified in the LNCaP/miR-331-3p microarray. ** indicates previously reported targets of miR-331-3p in PCa. (C) Box and Whisker plot analyses of PLCγ1, RALA, MARCKS, RRBP1 and miR-331-3p expression in patient tumor v non-malignant prostate tissue. A value of 1.0 is the calculated ratio where the expression of the miRNA or direct target was unchanged between tumor and NAT, and is indicated by the dotted line. (D) Luciferase reporter gene analysis of the 3’-UTR of putative miR-331-3p targets PLCγ1 and RALA, in C4-2B PCa cells transiently overexpressing miR-NC or miR-331-3p. (E) Luciferase reporter assays of the 3’-UTR of RALA with the two seed regions for miR-331-3p in the 3’-UTR mutated, in LNCaP and C4-2B PCa cells transiently overexpressing miR-NC or miR-331-3p. For all data shown, Error bars = SD; are representative of three independent experiments and *p<0.05, **p<0.005.