Subcellular Localization of p44/WDR77 Determines Proliferation and Differentiation of Prostate Epithelial Cells Shen Gao, Zhengxin Wang* Department of Cancer Biology, The University of Texas MD Anderson Cancer Center, Houston, Texas, United States of America Abstract The molecular mechanism that controls the proliferation and differentiation of prostate epithelial cells is currently unknown. We previously identified a 44-kDa protein (p44/wdr77) as an androgen receptor-interacting protein that regulates a set of androgen receptor target genes in prostate epithelial cells and prostate cancer. In this study, we found that p44 localizes in the cytoplasm of prostate epithelial cells at the early stage of prostate development when cells are proliferating, and its nuclear translocation is associated with cellular and functional differentiation in adult prostate tissue. We further demonstrated that cytoplasmic p44 protein is essential for proliferation of prostate epithelial cells, whereas nuclear p44 is required for cell differentiation and prostate- specific protein secretion. These studies suggest a novel mechanism by which proliferation and differentiation of prostate epithelial cells are controlled by p44’s location in the cell. Citation: Gao S, Wang Z (2012) Subcellular Localization of p44/WDR77 Determines Proliferation and Differentiation of Prostate Epithelial Cells. PLoS ONE 7(11): e49173. doi:10.1371/journal.pone.0049173 Editor: Daotai Nie, Southern Illinois University School of Medicine, United States of America Received August 2, 2012; Accepted October 8, 2012; Published November 8, 2012 Copyright: 2012 Gao, Wang. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was supported in part by the National Institutes of Health through MD Anderson’s Cancer Center Support Grant (CA016672) and the Institutional Research Grant Programs from MD Anderson Cancer Center. No additional external funding was received for this study. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: zhenwang@mdanderson.org Introduction During development of a multicellular organism, cells prolifer- ate for a defined length of time before they begin functional differentiation [1,2,3]. Precise regulation of terminal cell division is needed to ensure production of proper numbers of differentiated cells at the appropriate time. Development of the prostate gland is a dynamic process in which epithelial cells proliferate and then functionally differentiate [4,5]. Androgen signaling through the androgen receptor (AR) induces the growth of the prostate epithelium in early development [6] and is required later for the production of prostate-secreted proteins [7]. Other factors (NKX3.1, Hox13, FoxA, Shh, Sox9, Fgf7/10, Wnt5a, Bmp4/7, and Notch1) specifically affect ductal morphology, budding, and branching of epithelium during mouse prostate development [5]. However, the molecular mechanism that determines the timing of prostate epithelial proliferation and differentiation is currently unknown. After puberty, a man’s serum testosterone level peaks, and his prostate gland matures [8]. Serum testosterone levels then decrease with age by an average of 0.8% per year while the prostate continues to grow [9,10,11]. The age-related proliferation of prostate epithelial cells is a critical step leading to prostatic intraepithelial neoplasia (PIN) and prostate cancer [10,11,12,13,14,15]. Very little is known about what regulates this age- related growth of the prostate gland. Therefore, unraveling the molecular mechanism that controls prostate epithelial proliferation and differentiation is an important goal for understanding not only prostate development but also prostate tumorigenesis. To search for factors that regulate the AR functions, we previously purified and cloned a novel AR-interacting protein (p44) [16] that regulates expression of a subset of AR-target genes in the prostate and prostate cancer [16,17,18,19]. This protein is designated as WD40 Repeat Domain 77 (WDR77) in the NCBI Gene Bank (http://www.ncbi.nlm.nih. gov/homologene/?term = WDR77). Human p44 is composed of 342 amino acid residues and 7 putative WD-40 repeats. The protein sequence of p44 is identical to that of a component (MEP50) of the methylosome [20] and a subunit (WD45) of the SMN complex [21]. The methylosome complex contains PRMT5, MEP50, pICln, and Sm proteins and mediates the assembly of spliceosomal snRNP [20,22]. SMN, the protein involved in spinal muscular atrophy, is part of a complex containing the Sm proteins, WD45, and PRMT5. The SMN complex is necessary and sufficient for assembly of UsnRNA [23,24]. Although biochemically identified in the methylosome and SMN complex, the functional role of MEP50/WD45 in the splicing process has not been proven in vivo. We found the p44 immunostaining signal was strong in the nuclei of epithelial cells in normal prostate tissue but absent in the stromal cells [18]. In contrast, the nuclear staining in tumors is significantly lower, while immunostaining in the cytoplasm is strong. Similarly, p44 has been found to localize to the cytoplasm of prostate cancer cell lines LNCaP, 22RV1, PC3, and DU145 [25]. When p44 was selectively expressed in the nucleus by fusing a strong nuclear localization signal (NLS) to its N-terminus, the nuclear p44 strongly inhibited the growth of prostate cancer cells in tissue culture and in prostate tumors in nude mice by arresting PLOS ONE | www.plosone.org 1 November 2012 | Volume 7 | Issue 11 | e49173