Skip to main content
. 2013 Feb 5;288(13):9001–9010. doi: 10.1074/jbc.M112.437897

FIGURE 2.

FIGURE 2.

Secreted HSP90α induces fibronectin expression but represses E-cadherin, connexin-43, and connexin-26 levels in CRC cells. A, RT-PCR was performed to detect E-cadherin, fibronectin, connexin-43, and connexin-26 mRNA levels in HCT-115 and SW480 cells treated for 24 h with serum starvation CM in the presence of preimmune IgG or anti-HSP90α antibody. Representative data from three independent experiments are shown. The quantitative data were also obtained using real-time RT-PCR. Taking HCT-115 cells as an example, cellular E-cadherin, connexin-26, and connexin-43 mRNA levels were decreased to 13, 11, and 14%, respectively; however, fibronectin expression was increased to 790% in response to CM plus preimmune IgG. The levels of E-cadherin, connexin-26, connexin-43, and fibronectin mRNA expression could be recovered to 50, 91, 79, and 129%, respectively, when anti-HSP90α antibody was added instead, suggesting that the effects of CM were at least partly attributable to the action of HSP90α secreted in CM. CTRL, control. B, immunoblot analyses were performed to detect E-cadherin, fibronectin, connexin-43, and connexin-26 protein levels in HCT-115 and SW480 cells treated for 24 h with CM in the absence or presence of preimmune IgG or anti-HSP90α antibody. Representative data from three independent experiments are shown. C, RT-PCR was performed to analyze E-cadherin, fibronectin, connexin-43, and connexin-26 mRNA levels in HCT-115, LoVo, SW480, and SW620 cells treated for 24 h with 15 μg/ml rHSP90α. Representative images from three independent experiments are shown. The quantitative data were also obtained using real-time RT-PCR. Taking HCT-115 cells as an example, cellular levels of E-cadherin, connexin-26, and connexin-43 mRNAs were decreased to 9, 24, and 28%, respectively; however, fibronectin expression was increased by 4.2-fold in rHSP90α-treated cells compared with control cells treated with PBS. D, immunoblot analyses were performed to detect E-cadherin, fibronectin, connexin-43, and connexin-26 protein levels in HCT-115, LoVo, SW480, and SW620 cells treated for 24 h with PBS or 15 μg/ml rHSP90α. Representative data from three independent experiments are shown.