title: A proline rich acidic protein PRAP identified from uterine luminal fluid of estrous mice is able to enhance the estrogen responsiveness of Ishikawa cells.
abstract: Using mice as experimental animals, proteins in the uterine luminal fluid (ULF) from both adults and diethylstilbestrol dipropionate (DES)-treated immature animals were resolved by 2D gel electrophoresis. Two of the protein spots, (a) and (b) around the positions of 18-20 kDa, in the adult ULF were not found in the DES-treated ULF. Automated Edman degradation established the same N-terminal sequences of AHQVPVKTKGKHVFP for the two protein spots. Two trypsin digests of spot (a) were analyzed using CID MS/MS to establish the peptide sequences DNQLGPLLPEPK and RPDAMTWVETEDILSHLR. These partial sequences were confirmed in the cDNA-deduced mouse proline rich acidic protein (PRAP). Using human Ishikawa cell line as a surrogate endometrial model, we demonstrated rapid entrance of exogenous PRAP into the cells and its ability to enhance alkaline phosphatase activity of the E(2) -stimulated cells. Further, the transcripts of five estrogen-responsive genes, including ALPP (Placental alkaline phosphtase), ALPPL (placental alkaline phosphatase-like 2), TGF (transforming growth factor), PR (progesterone receptor), and Wnt7a, were measured after the cell incubation in modified Eagle medium containing 0.1 nM E(2) , or 0-25 µM PRAP, or both together at 37°C for 48 h. As compared with the control, E(2) alone increased the transcripts of ALPP, ALPPL, TGF-α, and PR, and reduced the transcript of Wnt7a, whereas PRAP alone had a slight impact on their expression. E(2) together with PRAP greatly increased the E(2) -stimulated transcriptions of ALPP, ALPPL, TGF-α, and PR, and markedly reduced the E(2) -suppressed transcription of Wnt7a.
authors.affiliation: Institute of Biochemical Sciences, College of Life Science, National Taiwan University, Taipei, Taiwan.
authors.forename: Tsai-Fu
authors.initials: TF
authors.lastname: Hsieh
authors.forename: Chia-Jen
authors.initials: CJ
authors.lastname: Tseng
authors.forename: Jyh-Bing
authors.initials: JB
authors.lastname: Tang
authors.forename: Yee-Hsiung
authors.initials: YH
authors.lastname: Chen
mesh_terms: D000328:Adult; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D017931:DNA Primers; D015180:Electrophoresis, Gel, Two-Dimensional; D004705:Endocytosis; D004967:Estrogens / Q000378:metabolism*; D004971:Estrus; D005260:Female; D006801:Humans; D051379:Mice; D008813:Mice, Inbred ICR; D008969:Molecular Sequence Data; D011392:Proline / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D053719:Tandem Mass Spectrometry; D014599:Uterus / Q000378:metabolism*
chemical_list: D017931:DNA Primers; D004967:Estrogens; D011506:Proteins; D011392:Proline