Mutant p53 regulates ovarian cancer transformed phenotypes through autocrine matrix deposition
Marcin P. Iwanicki, Hsing-Yu Chen, Claudia Iavarone, Ioannis K. Zervantonakis, Taru Muranen, Marián Novak, Tan A. Ince, Ronny Drapkin, Joan S. Brugge
Marcin P. Iwanicki, Hsing-Yu Chen, Claudia Iavarone, Ioannis K. Zervantonakis, Taru Muranen, Marián Novak, Tan A. Ince, Ronny Drapkin, Joan S. Brugge
View: Text | PDF
Research Article Cell biology Oncology

Mutant p53 regulates ovarian cancer transformed phenotypes through autocrine matrix deposition

Abstract

High-grade serous ovarian carcinoma (HGS-OvCa) harbors p53 mutations and can originate from the epithelial cell compartment of the fallopian tube fimbriae. From this site, neoplastic cells detach, survive in the peritoneal cavity, and form cellular clusters that intercalate into the mesothelium to form ovarian and peritoneal masses. To examine the contribution of mutant p53 to phenotypic alterations associated with HGS-OvCA, we developed live-cell microscopy assays that recapitulate these early events in cultured fallopian tube nonciliated epithelial (FNE) cells. Expression of stabilizing mutant variants of p53, but not depletion of endogenous wild-type p53, in FNE cells promoted survival and cell-cell aggregation under conditions of cell detachment, leading to the formation of cell clusters with mesothelium-intercalation capacity. Mutant p53R175H-induced phenotypes were dependent on fibronectin production, α5β1 fibronectin receptor engagement, and TWIST1 expression. These results indicate that FNE cells expressing stabilizing p53 mutants acquire anchorage independence and subsequent mesothelial intercalation capacity through a mechanism involving mesenchymal transition and matrix production. These findings provide important new insights into activities of mutant p53 in the cells of origin of HGS-OvCa.

Authors

Marcin P. Iwanicki, Hsing-Yu Chen, Claudia Iavarone, Ioannis K. Zervantonakis, Taru Muranen, Marián Novak, Tan A. Ince, Ronny Drapkin, Joan S. Brugge

×

Figure 2

Mutant p53R175H (m-p53R175H) promotes survival of detached fallopian tube nonciliated epithelial (FNE) cells independently of cell-cell adhesion.

Options: View larger image (or click on image) Download as PowerPoint
Mutant p53R175H (m-p53R175H) promotes survival of detached fallopian tub...
(A) H&E staining of disseminating carcinoma cell clusters from the fallopian tube (upper) and phase-contrast images of control and FNE-m-p53R175H cells cultured in suspension for 24 hours. Arrows point to detached carcinoma cells from the surface of fimbriae (upper), and to compacted FNE-m-p53R175H cells. Scale bar: 300 μm. (B) Representative (n = 8 videos) video clips of control and FNE-m-p53R175H cells cultured in suspension for the indicated times from Supplemental Video 3. Eight movies were recorded per cell type. Scale bar: 150 μm. (C) Representative time course of the percentage reduction in cell cluster area in suspension for control and FNE-m-p53R175H cells. n = 8 cell clusters scored per condition. Data shown as the mean ± SEM. The percentage reduction in cell area at the latest time point was measured in 3 independent experiments on n = 29 control and n = 31 FNE-m-p53R175H cellular clusters. Each cellular cluster contained 100–150 cells. (D) Upper panel: Schematic depiction of the microscopy-based detached-single-cell assay. Lower panel: Representative phase-contrast images of the outcomes observed following culture of single cells in suspension for 72 hours (Supplemental Video 4). Scale bar: 100 μm. (E) Quantification of single-cell death (upper panel) or proliferation (lower panel) in suspension for 72 hours. This experiment was repeated 3 times with n = 50–70 single cells analyzed per condition per experiment. Data shown as the median (horizontal bar), interquartile range (box), and minimum/maximum values (whiskers). Statistical analysis performed using 2-tailed Student’s t test. *P <0.05.