Suppression of tissue factor expression, cofactor activity, and metastatic potential of murine melanoma cells by the N‐terminal domain of adenovirus E1A 12S protein

Abstract

Tissue factor, the cellular initiator of blood coagulation, has been implicated as a determinant of metastatic potential in human melanoma cells. Here, we report that differential expression of tissue factor in murine melanoma cell lines of known metastatic behavior is mediated by AP-1-dependent and 12S E1A oncoprotein-repressible gene transcription. When compared to weakly metastatic C10 cells, highly metastatic M4 cells possessed elevated levels of tissue factor cofactor activity, transfected promoter activity, and heterodimeric AP-1 DNA-binding complexes containing Fra-1. Transient co-expression of the adenovirus E1A 12S oncoprotein strongly repressed transcription of an AP-1-driven tissue factor reporter gene indicating the additional requirement of N-terminal E1A-interacting coactivators. Stable expression of E1A mutants defective in CBP/p300-binding failed to suppress tissue factor expression and experimental metastasis by M4 cells while clones expressing wild type E1A exhibited greatly reduced tissue factor cofactor activity and metastatic potential in vivo. Overexpression of functional tissue factor in cells containing wild type E1A failed to restore the highly metastatic M4 phenotype suggesting that additional E1A-responsive and CBP/p300-dependent genes are required to facilitate metastasis of murine melanoma cells demonstrating high tissue factor expression and cofactor activity.

Ex vivo and In vivo Delivery of Anti-Tissue Factor Short Interfering RNA Inhibits Mouse Pulmonary Metastasis of B16 Melanoma Cells

“…In preliminary experiments, the M4 cells exhibited a higher metastatic tendency than B16 cells, coinciding with a 2-to 2.5-fold higher relative (normalized to GAPDH) expression of tissue factor mRNA compared with B16 cells (data not shown). These data were consistent with the previously reported correlation of the metastatic tendency of melanoma cells with their levels of tissue factor expression (8,10). Although the M4 cells displayed a higher metastatic tendency than B16 cells, the former proved relatively refractory to liposome-mediated transfection under standard transfection conditions (only 40-50% knockdown in cell culture; data not shown).…”

“…More recently, tissue factor has also been implicated in various processes promoting cancer development and spreading. Many studies have reported that tissue factor plays a major role in cancer-driven angiogenesis (3 -6), whereas tissue factor expression seems to correlate with progression/growth, invasion, and metastasis of various types of cancer cells (3,4,(7)(8)(9)(10). Tissue factor expression may thus be a clinically useful prognostic factor (7).…”

Endothelial cell expression of tissue factor in sickle mice is augmented by hypoxia/reoxygenation and inhibited by lovastatin

“…22 Polyclonal antibodies of the immunoglobulin G (IgG) class were enriched from rabbit antisera by ammonium sulfate fractionation. For simplicity, we will refer to this partially purified antibody preparation as "␣TF."…”

Endothelial nitric oxide synthase and nitric oxide regulate endothelial tissue factor expression in vivo in the sickle transgenic mouse

“…We measured expression of TF by the pulmonary vein endothelium exactly as we previously described [3], using immunofluorescence microscopic review of tissue sections triple stained for nuclei and endothelial marker CD31 and murine TF, with fluorescein isothiocyanate and tetramethylrhodamine-5(and6)-isothiocyanate-labeled secondary antibodies. For this, we used the partially purified polyclonal antibody preparation to murine TF [36] that we previously described [3]. Quality control measures and negative controls were as previously detailed [3].…”