Cancer-Stem-Cell-Like, Wnt/TCF Responsive Cells Are Activated by Pax8 PPAR? Fusion Protein.

Abstract

Pax8 PPARγ Fusion Protein (PPFP) occurs in ~35% of follicular thyroid carcinoma cases. Expression of PPFP in the non-transformed rat thyroid cell line PCCL3 conferred on the cells the ability to invade through matrigel and to form colonies in anchorage independent conditions.

PPFP also increased the percentage of cells that have activated β-catenin/TCF. We transduced control and PPFP-expressing PCCL3 thyroid cell lines with two different β-catenin/TCF-GFP reporter systems. PPFP expressing cell lines contained more than twice the percent of GFP positive cells compared to control cell lines.

A hierarchy existed within the cell lines based on the TCF activation status. A single TCF_GFP+ cell generated a clonal population containing both GFP+ and GFP- cells, whereas clonal populations derived TCF_GFP- cells remained GFP-. Single cells sorted by TCF status demonstrated that very few TCF_GFP- cells can generate any TCF_GFP+ cells.

The TCF responsive cells exhibited increased proliferative potential and invasive capacity. GFP+ cells were twice as enriched for anchorage independent colony forming cells than GFP cells. More strikingly, GFP positive clones were 5-10 times more invasive than negative clones, which exhibited the same invasive potential as control cells.

The hierarchy and more transformed phenotype of TCF responsive cells indicate that they have the in vitro properties of cancer stem cells.

Full agonists of PPARγ further increased the effects of PPFP. The β-catenin/TCF population in PPFP expressing cells increased again while similarly treated empty vector control cells were unaffected. Invasion was also increased by the PPARγ agonists. The effects were blocked by PPARγ antagonists, demonstrating PPARγ specificity. Selective PPARγ agonists which only activate a fraction of the PPARγ effects also increased the effects of PPFP. We could then assume that the adipogenic pathways activated by full agonists but not selective PPARγ agonists did not contribute to PPFP oncogenesis.

These data suggested that PPFP acts via its PPARγ domains to expand the Wnt/TCF active cell fraction and that these cells have the properties of cancer stem cells. The pathways by which PPFP effect oncogenesis are activated by both full and partial PPARγ agonists.