Research Project:
NUTRITION, AGING, AND VISUAL FUNCTION
Location: Human Nutrition Research Center on Aging
Title: Proteasome-dependent regulation of signal transduction in retinal pigment epithelial cells
Authors
![item](https://webarchive.library.unt.edu/eot2008/20090514204258im_/http://www.ars.usda.gov/incme/images/bullet.gif) | Fernandes, Alexandre - HNRCA AT TUFTS | ![item](https://webarchive.library.unt.edu/eot2008/20090514204258im_/http://www.ars.usda.gov/incme/images/bullet.gif) | Guo, Weimin - HNRCA AT TUFTS | ![item](https://webarchive.library.unt.edu/eot2008/20090514204258im_/http://www.ars.usda.gov/incme/images/bullet.gif) | Zhang, Xinyu - HNRCA AT TUFTS | ![item](https://webarchive.library.unt.edu/eot2008/20090514204258im_/http://www.ars.usda.gov/incme/images/bullet.gif) | Gallagher, Matthew - HNRCA AT TUFTS | ![item](https://webarchive.library.unt.edu/eot2008/20090514204258im_/http://www.ars.usda.gov/incme/images/bullet.gif) | Mircea, Ivan - HNRCA AT TUFTS | ![item](https://webarchive.library.unt.edu/eot2008/20090514204258im_/http://www.ars.usda.gov/incme/images/bullet.gif) |
Taylor, Allen
| ![item](https://webarchive.library.unt.edu/eot2008/20090514204258im_/http://www.ars.usda.gov/incme/images/bullet.gif) | Pereira, Paulo - HNRCA AT TUFTS | ![item](https://webarchive.library.unt.edu/eot2008/20090514204258im_/http://www.ars.usda.gov/incme/images/bullet.gif) | Shang, Fu - HNRCA AT TUFTS |
Submitted to: Experimental Eye Research
Publication Type:
Peer Reviewed Journal
Publication Acceptance Date: July 31, 2006
Publication Date: October 5, 2006
Citation: Fernandes, A.F., Guo, W., Zhang, X., Gallagher, M., Mircea, I., Taylor, A., Pereira, P., Shang, F. 2006. Proteasome-dependent regulation of signal transduction in retinal pigment epithelial cells. Experimental Eye Research. 83(6):1472-81.
Interpretive Summary: Retinal pigment epithelial (RPE) cells have an active ubiquitin-proteasome pathway (UPP), yet the function of the UPP in RPE still needs to be clarified. This study aims to determine the role of UPP in controlling the levels and activities of transcription factors, hypoxia-inducible factor (HIF) and NF-KappaB. The UPP was inhibited with proteasome-specific inhibitors to determine the activation of HIF and NF-kappaB in addition to the expression and secretion of pro-angiogenic factors. Results showed that HIF-1alpha was not found in ARPE-19 cells under normal culture conditions, yet when proteasome activity was inhibited, HIF-1alpha accumulated in RPE cells based on a specific timeframe. Protease inhibition also led to an increase of HIF-1alpha, levels of mRNA for vascular endothelial growth factor (VEGF), and angiopoietin-2 (Ang-2) in RPE cells, which were up to 7 times higher than usual. In addition, proteasome inhibition was also associated with a 2-fold increase in mRNA levels for angiopoietin-1 (Ang-1) as well as VEGF secretion in comparison to its normal culture conditions. On the other hand, proteasome inhibition reduced NF-kappaB activation, while the expression and secretion of monocyte chemoattractant protein-1 (MCP-1) were notably weakened by RPE cells. The data suggests that UPP plays a significant role in regulating HIF and NF-kappaB in RPE cells. Consequences of UPP impairment includes accumulation of HIF-1alpha and diminished NF-kappaB activation; enhanced expression and secretion of pro-angiogenic factors; and weakened expression of MCP-1 could also develop. Overall, the results indicate that impairment of UPP could lead to the development of AMD-related phenotypes.
Technical Abstract: As in many other types of cells, retinal pigment epithelial (RPE) cells have an active ubiquitin-proteasome pathway (UPP). However, the function of the UPP in RPE remains to be elucidated. The objective of this study is to determine the role of the UPP in controlling the levels and activities of transcription factors hypoxia-inducible factor (HIF) and NF-kappaB. We inhibited the UPP with proteasome-specific inhibitors and determined the activation of HIF and NF-kappaB as well as the expression and secretion of pro-angiogenic factors. HIF-1alpha was not detectable in ARPE-19 cells under normal culture conditions. However, when proteasome activity was inhibited, HIF-1alpha accumulated in RPE in a time-dependent manner. Consistent with accumulation of HIF-1alpha in the cells, levels of mRNA for vascular endothelial growth factor (VEGF) and angiopoietin-2 (Ang-2) in RPE were up to 7-fold higher upon inhibition of the proteasome. Proteasome inhibition was also associated with a 2-fold increase in levels of mRNA for angiopoietin-1 (Ang-1). ARPE-19 cells secrete significant levels of VEGF under normal culture conditions. Inhibition of proteasome activity increased the secretion of VEGF by 2-fold. In contrast to the increase in HIF activity, NF-kappaB activation was reduced by proteasome inhibition. In addition, the expression and secretion of monocyte chemoattractant protein-1 (MCP-1) by RPE were substantially attenuated by the inhibition of proteasome activity. These data demonstrate that the UPP plays an important role in modulating the activities of HIF and NF-kappaB in the RPE. Consequences of an impairment of the UPP include accumulation of HIF-1alpha and diminished NF-kappaB activation, which lead to enhanced expression and secretion of pro-angiogenic factors and attenuated expression of MCP-1. Taken together, these data predict that the impairment of the UPP could lead to the development of AMD-related phenotypes.
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