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Qian Chen, Yusuke Takahashi, Kyungwon Lee, Kazuhiro Oka, Jian-Xing Ma; Differential Roles of Splice Variants of Very Low-density Lipoprotein Receptor in Regulating the Wnt Signaling Pathway. Invest. Ophthalmol. Vis. Sci. 2014;55(13):6011.
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© ARVO (1962-2015); The Authors (2016-present)
Very low-density lipoprotein receptor (VLDLR) is a multi-ligand receptor and has diverse roles beyond lipoprotein metabolism. Our previous studies have shown that VLDLR negatively regulates Wnt signaling in the retina. Two alternative splice variants of VLDLR, variant I (VLDLRI) and variant II (VLDLRII) are differentially expressed in tissues. However, the significance of distinct tissue distribution and the differential effects of these variants have not been investigated. The purpose of this study is to investigate the differential roles of splice variants of VLDLR in regulating Wnt signaling.
The expression of VLDLRI and VLDLRII in various tissues of mouse (brain, retina, heart, liver, kidney, spleen, ovary, uterus, muscle, adipose tissue) was analyzed by RT-PCR. Levels of the shed VLDLR extracellular domain and full-length VLDLR were evaluated by Western blot analysis in the culture medium or cell pellet of CHO cells expressing either VLDLRI or VLDLRII, and the ratio of shed/full-length VLDLRI or VLDLRII were determined by densitometry. The impact of shed/full-length VLDLRI and VLDLRII on Wnt signaling activity was evaluated by a luciferase-based assay using ARPE-19 and hTERT stable TOP-FLASH cell lines.
Most of the examined murine tissues expressed both VLDLRI and VLDLRII, while the heart solely expressed VLDLRI, and the retina exclusively expressed VLDLRII. The exclusive expression of VLDLRII was also verified in the human retina. A shed VLDLR extracellular domain was detected in conditioned medium from cells expressing full-length VLDLRI or VLDLRII. The ratio of shed/full-length VLDLR was dramatically higher in cells expressing VLDLRII than VLDLRI. Full-length VLDLRI and VLDLRII exhibited similar inhibitory effects on Wnt signaling activities. However, the shedding of VLDLR in conditioned medium showed more potent effects than full-length counterparts on inhibiting Wnt signaling activities. Furthermore, conditioned medium from cells expressing VLDLRII showed a more potent inhibitory effect on Wnt signaling than that from cells expressing VLDLRI.
The retina expressed VLDLRII exclusively and had more repaid shedding of the extracellular domain, which showed a more potent regulatory effect on Wnt signaling than VLDLRI. The shed VLDLR extracellular domain may represent a novel mechanism for Wnt signaling regulation in the retina.
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