title: Poly r(C) binding protein-1 is central to maintenance of cancer stem cells in prostate cancer cells.
abstract: OBJECTIVE
  To investigate global proteomic changes induced in CD44+CD24- stem cells isolated from the prostate cancer cell lines, LNCaP and DU145, post prolonged TGF-β treatment in order to understand underlying mechanisms that promote stemness in prostate cancer cells.
  
  
  METHODS
  CD44+CD133+α2β1Integrin+CD24- population was isolated from mock or TGF-β treated (7 days) prostate cancer cell line, LNCaP, through fluorescent activated cell sorting. Cell lysates were obtained from the ±TGF-β cell population and proteomics profiling (MS/MS) was performed by mass spectrometry. Relative enrichment or depletion in the CD44+CD24-population post-TGF-β treatment was determined relative to mock-treated CD44+CD24- cells post normalization to GAPDH expression levels. RESULTS obtained from MS/MS were validated using immunoblotting. Functional validation of one putative regulator was performed using gain-of-function strategy to investigate its role in rendering stemness in LNCaP and DU145 cells in vitro and in promoting tumorigenicity in vivo.
  
  
  RESULTS
  TGF-β treatment caused significant enrichment of CD44+CD24- population in LNCaP cells (22.35 ± 0.94% in mock treated vs 95.23 ± 2.34% in TGF-β treated cells; P < 0.01), which were also positive for CD133 and α2β1Integrin. Mass spectrometry analysis of the enriched cell population revealed that sixty-three proteins were either up- or down-regulated greater than five folds, out of which the poly r(C) binding protein (PCBP)-1 was the most down-regulated (9.31 ± 0.05 folds). Ectopic overexpression of PCBP1 in LNCaP and DU145 cells not only attenuated enrichment of CD44+CD133+CD24- population in these cells following TGF-β treatment, but also significantly decreased tumorigenicity of the stem cell subset, as assessed by in vitro soft agar colony formation and in vivo xenograft assays.
  
  
  CONCLUSIONS
  Our proteomic profiling and subsequent validation indicate that PCBP1 is central to CSCs enrichment and functionality in prostate cancer.
authors: Department of Urology and Andrology, Ninth People's Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai, China.
authors: Qi
authors: Q
authors: Chen
authors: Zhi-kang
authors: ZK
authors: Cai
authors: Yan-bo
authors: YB
authors: Chen
authors: Meng
authors: M
authors: Gu
authors: Da-chao
authors: DC
authors: Zheng
authors: Juan
authors: J
authors: Zhou
authors: Zhong
authors: Z
authors: Wang
mesh_terms: D045744:Cell Line, Tumor; D002469:Cell Separation; D004268:DNA-Binding Proteins; D034441:Heterogeneous-Nuclear Ribonucleoproteins / Q000096:biosynthesis*; D006801:Humans; D008297:Male; D014411:Neoplastic Stem Cells / Q000378:metabolism*; D011471:Prostatic Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D040901:Proteomics; D016601:RNA-Binding Proteins; D053719:Tandem Mass Spectrometry; D016212:Transforming Growth Factor beta / Q000008:administration & dosage / Q000378:metabolism
chemical_list: D004268:DNA-Binding Proteins; D034441:Heterogeneous-Nuclear Ribonucleoproteins; C094604:PCBP1 protein, human; D016601:RNA-Binding Proteins; D016212:Transforming Growth Factor beta