[Show abstract][Hide abstract] ABSTRACT: The splicing factor SPF45 (RBM17) is frequently overexpressed in many solid tumors, and stable expression in HeLa cells confers resistance to doxorubicin and vincristine. In this study, we characterized stable transfectants of A2780 ovarian carcinoma cells. In a 3-day cytotoxicity assay, human SPF45 overexpression conferred 3- to 21-fold resistance to carboplatin, vinorelbine, doxorubicin, etoposide, mitoxantrone, and vincristine. In addition, resistance to gemcitabine and pemetrexed was observed at the highest drug concentrations tested. Knockdown of SPF45 in parental A2780 cells using a hammerhead ribozyme sensitized A2780 cells to etoposide by approximately 5-fold relative to a catalytically inactive ribozyme control and untransfected cells, suggesting a role for SPF45 in intrinsic resistance to some drugs. A2780-SPF45 cells accumulated similar levels of doxorubicin as vector-transfected and parental A2780 cells, indicating that drug resistance is not due to differences in drug accumulation. Efforts to identify small molecules that could block SPF45-mediated drug resistance revealed that the selective estrogen receptor (ER) modulators tamoxifen and LY117018 (a raloxifene analogue) partially reversed SPF45-mediated drug resistance to mitoxantrone in A2780-SPF45 cells from 21-fold to 8- and 5-fold, respectively, but did not significantly affect the mitoxantrone sensitivity of vector control cells. Quantitative PCR showed that ERbeta but not ERalpha was expressed in A2780 transfectants. Coimmunoprecipitation experiments suggest that SPF45 and ERbeta physically interact in vivo. Thus, SPF45-mediated drug resistance in A2780 cells may result in part from effects of SPF45 on the transcription or alternate splicing of ERbeta-regulated genes.
[Show abstract][Hide abstract] ABSTRACT: Purified ribonucleoprotein complexes of human parainfluenza virus type 3 (HPIV-3) virions required, in addition to the viral proteins, soluble cytoplasmic proteins from uninfected cells for the synthesis of mRNAs in vitro. In contrast to Sendai virus transcription, in vitro RNA synthesis from HPIV-3 ribonucleoprotein complexes was not stimulated significantly by purified tubulin. Moreover, cytoplasmic extract depleted of tubulin by immunoprecipitation stimulated HPIV-3 transcription effectively, suggesting involvement of a host protein(s) other than tubulin in the HPIV-3 transcription process. The transcription stimulatory factor was purified from uninfected cell extract by conventional chromatography and was found to contain a major 43-kDa polypeptide. In Western blot (immunoblot) analysis, this protein reacted with antiactin antibody, suggesting that the 43-kDa polypeptide is actin. This possibility was further supported by its polymerization activity and properties of binding to blue-Sepharose and heparin-Sepharose columns. Furthermore, when the cell extract was depleted of actin by immunoprecipitation by antiactin antibody, the stimulatory activity was abolished, indicating an involvement of actin in the stimulation of HPIV-3 transcription. After purification from RNAses, similar stimulatory activity associated with the 43-kDa protein was detected in other cell lines as well, including CV-1, HeLa, and BHK.
Full-text · Article · Jul 1991 · Journal of Virology
[Show abstract][Hide abstract] ABSTRACT: 1. Catalase activity was partially purified from body wall muscle of the parasitic nematode, Ascaris suum, and was similar to catalases isolated from mammalian tissues. It exhibited a broad pH optimum and was unaffected by 2 mM ethylenediaminetetra-acetate. In contrast, it was inhibited reversibly by 1 mM cyanide and irreversibly by prior incubation in 40 mM 3-amino-1:2:4-triazole for 1 hr or heating at 80 degrees C for 15 min. 2. Catalase activity was highest in the unembryonated "egg" and decreased dramatically as development proceeded. 3. Catalase activity in adult body wall muscle was similar to that in rat skeletal muscle, but dramatically lower than that in rat liver. Catalase activity was barely detectable in A. suum testis. 4. Cytochrome-c peroxidase activity did not appear to be present in adult A. suum muscle mitochondria.
No preview · Article · Feb 1990 · Comparative biochemistry and physiology. B, Comparative biochemistry
[Show abstract][Hide abstract] ABSTRACT: Typescript. Thesis (M.S.)--University of Toledo. "A thesis [submitted] as partial fulfillment of the requirements of the Master of Science degree in Biology." Includes bibliographical references (leaves 40-45).