Loss of CD73-mediated actin polymerization promotes endometrial tumor progression
Jessica L. Bowser, … , Kenneth Dunner Jr., Russell R. Broaddus
Jessica L. Bowser, … , Kenneth Dunner Jr., Russell R. Broaddus
Published December 7, 2015
Citation Information: J Clin Invest. 2016;126(1):220-238. https://doi.org/10.1172/JCI79380.
View: Text | PDF
Research Article Oncology

Loss of CD73-mediated actin polymerization promotes endometrial tumor progression

Abstract

Ecto-5′-nucleotidase (CD73) is central to the generation of extracellular adenosine. Previous studies have highlighted a detrimental role for extracellular adenosine in cancer, as it dampens T cell–mediated immune responses. Here, we determined that, in contrast to other cancers, CD73 is markedly downregulated in poorly differentiated and advanced-stage endometrial carcinoma compared with levels in normal endometrium and low-grade tumors. In murine models, CD73 deficiency led to a loss of endometrial epithelial barrier function, and pharmacological CD73 inhibition increased in vitro migration and invasion of endometrial carcinoma cells. Given that CD73-generated adenosine is central to regulating tissue protection and physiology in normal tissues, we hypothesized that CD73-generated adenosine in endometrial carcinoma induces an innate reflex to protect epithelial integrity. CD73 associated with cell-cell contacts, filopodia, and membrane zippers, indicative of involvement in cell-cell adhesion and actin polymerization–dependent processes. We determined that CD73-generated adenosine induces cortical actin polymerization via adenosine A1 receptor (A1R) induction of a Rho GTPase CDC42–dependent conformational change of the actin-related proteins 2 and 3 (ARP2/3) actin polymerization complex member N-WASP. Cortical F-actin elevation increased membrane E-cadherin, β-catenin, and Na+K+ ATPase. Together, these findings reveal that CD73-generated adenosine promotes epithelial integrity and suggest why loss of CD73 in endometrial cancer allows for tumor progression. Moreover, our data indicate that the role of CD73 in cancer is more complex than previously described.

Authors

Jessica L. Bowser, Michael R. Blackburn, Gregory L. Shipley, Jose G. Molina, Kenneth Dunner Jr., Russell R. Broaddus

×

Figure 6

The CD73 inhibitor AoPCP decreases circumferential cortical F-actin.

Options: View larger image (or click on image) Download as PowerPoint
The CD73 inhibitor AoPCP decreases circumferential cortical F-actin. (A)...
(A) Representative immunoblot of total actin (G- and F-actin) isolated from confluent HEC-1-A cells untreated (control) or treated with 50 μM 5′-AMP or 50 μM 5′-AMP plus 100 μM AoPCP for 50 minutes following 36 hours of hypoxia (1% O2, 5% CO2). DCF (10 μM) was included with treatments and controls. HEC-1-A cells were pretreated with 100 μM AoPCP or fresh Opti-MEM for 1 hour. 5′-AMP plus AoPCP no. 1 and 5′-AMP plus AoPCP no. 2 were independent replicates in a single experiment. Graph shows densitometric data from 6 independent experiments; 1 experiment (n = 7) was excluded because of values that exceeded ± 2 SD. **P < 0.005, *P < 0.05, and #P < 0.05; 1-way ANOVA with Tukey’s post test. 5′-AMP and AoPCP dose responses are shown in Supplemental Figures 8 and 9. (B) G-actin/F-actin ratio. **P < 0.005 and *P < 0.05; 1-way ANOVA with Holm-Sidak post test. (C) Alexa Fluor–conjugated phalloidin immunofluorescence in HEC-1-A cells. Cortical F-actin decreased with AoPCP. Scale bars: 20 μm. (D) F-actin intensity was assessed in at least 10 or more images (original magnification, ×20) using cellSens Dimension software, set at “Automatic Threshold.” **P < 0.001; 1-way ANOVA with Tukey’s post test. All data are expressed as the mean ± SEM.