title: The cancer/testis antigen prostate-associated gene 4 (PAGE4) is a highly intrinsically disordered protein.
abstract: The cancer/testis antigens (CTAs) are an important group of heterogeneous proteins that are predominantly expressed in the testis in the normal human adult but are aberrantly expressed in several types of cancers. Prostate-associated gene 4 (PAGE4) is a member of the CT-X family of CTAs that in addition to testis, is highly expressed in the fetal prostate, and may also play an important role both in benign and malignant prostate diseases. However, the function of this gene remains poorly understood. Here, we show that PAGE4 is a highly (100%) intrinsically disordered protein (IDP). The primary protein sequence conforms to the features of a typical IDP sequence and the secondary structure prediction algorithm metaPrDOS strongly supported this prediction. Furthermore, SDS-gel electrophoresis and analytical size exclusion chromatography of the recombinant protein revealed an anomalous behavior characteristic of IDPs. UV circular dichroism (CD) and NMR spectroscopy confirmed that PAGE4 is indeed a highly disordered protein. In further bioinformatic analysis, the PredictNLS algorithm uncovered a potential nuclear localization signal, whereas the algorithm DBS-Pred returned a 99.1% probability that PAGE4 is a DNA-binding protein. Consistent with this prediction, biochemical experiments showed that PAGE4 preferentially binds a GC-rich sequence. Silencing PAGE4 expression induced cell death via apoptosis and in mice carrying PCa xenografts, siRNA-mediated knockdown of the PAGE4 mRNA attenuated tumor growth in vivo. Furthermore, overexpressing PAGE4 protected cells from stress-induced death. To our knowledge, PAGE4 is the first example of a CTA that is an IDP with an anti-apoptotic function.
authors.affiliation: Department of Urology, James Buchanan Brady Urological Institute & the George O'Brien Urology Research Center, Johns Hopkins University School of Medicine, Baltimore, Maryland 21287, USA.
authors.forename: Yu
authors.initials: Y
authors.lastname: Zeng
authors.forename: Yanan
authors.initials: Y
authors.lastname: He
authors.forename: Fan
authors.initials: F
authors.lastname: Yang
authors.forename: Steven M
authors.initials: SM
authors.lastname: Mooney
authors.forename: Robert H
authors.initials: RH
authors.lastname: Getzenberg
authors.forename: John
authors.initials: J
authors.lastname: Orban
authors.forename: Prakash
authors.initials: P
authors.lastname: Kulkarni
mesh_terms: D000818:Animals; D000951:Antigens, Neoplasm / Q000737:chemistry* / Q000502:physiology; D017209:Apoptosis; D045744:Cell Line, Tumor; D002942:Circular Dichroism; D006801:Humans; D008297:Male; D051379:Mice; D016513:Mice, SCID; D009368:Neoplasm Transplantation; D011471:Prostatic Neoplasms / Q000473:pathology; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D034741:RNA, Small Interfering / Q000378:metabolism; D013737:Testis / Q000378:metabolism
chemical_list: D000951:Antigens, Neoplasm; C474505:PAGE4 protein, human; D034741:RNA, Small Interfering