Up-regulation of dicer, a component of the MicroRNA machinery, in prostate adenocarcinoma

Abstract

MicroRNAs are small noncoding 18- to 24-nt RNAs that are predicted to regulate expression of as many as 30% of protein-encoding genes. In prostate adenocarcinoma, 39 microRNAs are up-regulated, and six microRNAs are down-regulated. Production and function of microRNA requires coordinated processing by proteins of the microRNA machinery. Dicer, an RNase III endonuclease, is an essential component of the microRNA machinery. From a gene array analysis of 16 normal prostate tissue samples, 64 organ-confined, and four metastatic prostate adenocarcinomas, we identified an up-regulation of major components of the microRNA machinery, including Dicer, in metastatic prostate adenocarcinoma. Immunohistochemical studies on a tissue microarray consisting of 232 prostate specimens confirmed up-regulation of Dicer in prostatic intraepithelial neoplasia and in 81% of prostate adenocarcinoma. The increased Dicer level in prostate adenocarcinoma correlated with clinical stage, lymph node status, and Gleason score. Western blot analysis of benign and neoplastic prostate cell lines further confirmed Dicer up-regulation in prostate adenocarcinoma. Dicer up-regulation may explain an almost global increase of microRNA expression in prostate adenocarcinoma. The presence of up-regulated microRNA machinery may predict the susceptibility of prostate adenocarcinoma to RNA interference-based therapy.

Figure 1

Up-regulation of the components of the miR machinery in prostate cancer. A: Schematic representation of miR maturation. Up-regulated genes are underlined. B: Up-regulation of genes encoding miR machinery components in metastatic prostate adenocarcinoma. The fold change over organ-confined prostate adenocarcinoma is shown. POLR2A, polymerase (RNA) II polypeptide A; XPO5, exportin 5; EIF2C2, eukaryotic translation initiation factor 2C (Ago2); HSPCA, heat shock 90-kd protein 1, alpha; TNRC6B, trinucleotide repeat containing 6B; MOV10, Moloney leukemia virus 10, homolog; EIF2C1, eukaryotic translation initiation factor 2C1 (Ago1). C: Dicer mRNA and EIF2C2 mRNA levels in organ-confined and metastatic PCa (P < 0.05) assessed by gene array. Representative cases are shown.

Figure 2

Expression of Dicer in benign prostate epithelium. A, B: Basal cells express a high level of Dicer, whereas luminal cells are negative, as determined by immunohistochemistry; original magnification: ×200 (A) and ×400 (B). C: Dicer highlights basal cell hyperplasia, as determined by immunohistochemistry; original magnification, ×400.

Figure 3

Expression of Dicer in neoplastic prostate tissue. A: The largest benign gland in the center of the field shows Dicer immunoreactivity limited to the basal cells; glands in the right and left lower corners are affected by PIN, characterized by multilayered high-columnar cells with enlarged nuclei, prominent nucleoli, and higher cytoplasmic levels of Dicer, shown by immunohistochemistry; original magnification, ×200. B: Prostatic intraepithelial neoplasia examined by Dicer immunohistochemistry; original magnification, ×400. C: Prostate adenocarcinoma with smaller, haphazardly arranged glands of varying size, with high Dicer immunoreactivity, shown by immunohistochemistry; original magnification, ×200. D: Strong Dicer immunoreactivity in neoplastic cells of prostate adenocarcinoma, immunohistochemistry; original magnification, ×400.

Figure 4

Schematic representation of mean Dicer immunoreactivity in benign and malignant prostate tissue. A: Mean Dicer immunoreactivity in benign prostate, PIN, and PCa (P = 0.001). B: Mean Dicer immunoreactivity distinguishes PCa by stage (P = 0.001). C: Mean Dicer immunoreactivity is higher in N1 PCa (N0, no positive regional lymph nodes; N1, metastases in regional nodes) (P = 0.006). D: Mean Dicer immunoreactivity is higher in PCa with Gleason score >7 (P = 0.025).

Figure 5

Western blot and immunocytochemical analysis of Dicer expression in DU145 and RWPE-1 prostate cancer cell lines. A: Expression of Dicer in RWPE-1 and DU145 cell lines. DU145 cell line demonstrates higher level of Dicer by Western blot. When 70 μg of total protein extract were loaded, Dicer was detected in RWPE-1 cells but at a level dramatically lower than in DU145. B: Immunocytochemistry of Dicer in DU145 cell line; original magnification, ×400.