cDNA array analysis of SPARC-modulated changes in glioma gene expression.
We have demonstrated that secreted protein acidic and rich in cysteine (SPARC) is highly expressed in human gliomas and it promotes glioma invasion and delays tumor growth in vitro and in vivo. cDNA array analyses were performed to determine whether SPARC, which interacts at the cell surface, has an impact on intracellular signaling and downstream gene expression changes, which might account for some of its effects on invasion and growth. Using a doxycycline (dox)-controlled gene expression system, two cDNA array analyses were performed using a parental U87T2 clone (-SPARC) transfected with the dox-controlled transactivator and a U87T2 parental-derived SPARC-transfected clone, A2b2 (+SPARC). Array analysis performed between the parental and the SPARC-transfected clone (-dox) identified 13 upregulated genes and 14 downregulated genes. With the exception of PAI-1 and MMP2, the identified genes are novel with respect to SPARC's mechanism of action. Array analysis performed using the SPARC-transfected clone ( +/- dox) identified 2 types of gene regulation; one reversible upon SPARC suppression, the other irreversible. Two of the SPARC-induced genes, BIGH3 (irreversible by dox) and PAI-1 (reversible by dox) were further studied in additional SPARC-transfected clones, human astrocytoma tissues, and human glioma cell lines by RT-PCR and Northern blot analyses. The results indicate that: (1) the array results were validated, (2) the dox regulation was validated, and (3) the differential expression identified by the array analyses was present between normal brain and in human astrocytoma tissues and cell lines. Therefore, we conclude that these cDNA array analyses provide candidate genes involved in SPARC-mediated effects on glioma cell cycle progression, signaling, and migration, and that SPARC may induce reversible and irreversible gene expression changes. Further investigation of these candidates may shed insights into SPARC's role in glioma cell proliferation and invasion, and potential use as a therapeutic target.
Golembieski
William A
WA
Barbara Jane Levy Laboratory of Molecular Neuro-Oncology, Hermelin Brain Tumor Center, Department of Neurosurgery, Henry Ford Health Sciences Center, Detroit, MI 48202, USA.
Rempel
Sandra A
SA
D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001253:Astrocytes / Q000378:metabolism; D001254:Astrocytoma / Q000235:genetics; D015152:Blotting, Northern; D001921:Brain / Q000378:metabolism; D002471:Cell Transformation, Neoplastic / Q000235:genetics; D004318:Doxycycline / Q000494:pharmacology; D016326:Extracellular Matrix Proteins; D005333:Fetus; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic; D005910:Glioma / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D009361:Neoplasm Invasiveness / Q000235:genetics; D009363:Neoplasm Proteins / Q000235:genetics; D020411:Oligonucleotide Array Sequence Analysis; D015676:Osteonectin / Q000235:genetics / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D016212:Transforming Growth Factor beta; D014407:Tumor Cells, Cultured; D000099222:betaIG-H3 Protein
D000900:Anti-Bacterial Agents; D004318:Doxycycline; D016326:Extracellular Matrix Proteins; D009363:Neoplasm Proteins; D015676:Osteonectin; D016601:RNA-Binding Proteins; D016212:Transforming Growth Factor beta; D000099222:betaIG-H3 Protein; C428129:SERBP1 protein, human