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Yu-Chieh Wu, Meifang Zhu, Danielle M. Robertson; IGF-1R/INSR Hybrid in Proliferating Corneal Epithelial Cells. Invest. Ophthalmol. Vis. Sci. 2012;53(14):1826.
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Type I insulin-like growth factor receptor (IGF-1R) and insulin receptor (INSR) are highly homologous molecules, which heterodimerize to form IGF-1R/INSR hybrid (Hybrid-R). The nuclear localization of IGF-1R was recently shown in human corneal epithelial cells; however, the function of nuclear IGF-1R is not generally clear and the presence of the Hybrid-R is unknown. The purpose of this study is to characterize the role of IGF-1/IGF-1R and Hybrid-R signaling and the presence and biological significance of Hybrid-R in cultured human corneal epithelial cells.
IGF-1-mediated cell growth and signaling were examined in a human telomerized corneal epithelial (hTCEpi) cell line using reducing immunoprecipitation, immunoblotting and cell proliferation assays. The presence of Hybrid-R in hTCEpi and primary cultured human corneal epithelial cells was confirmed by immunofluorescence and reciprocal immunoprecipitation of whole cell lysates. The nuclear profile of IGF-1R/IGF-1R homodimer and Hybrid-R was determined in nuclear extracts of hTCEpi cells using immunoprecipitation with anti-IGF-1R (αIR3) and anti-INSR, followed by immunoblots with anti-IGF-1R. Chromatin-immunoprecipitation (ChIP)-sequencing with anti-IGF-1R and anti-INSR was used to identify potential genomic targets in hTCEpi cells.
In cultured hTCEpi cells, IGF-1 stimulated Akt signaling and promoted cell growth through IGF-1R activation; insulin at the same concentration did not show a significant effect. Reciprocal immunoprecipitation with antibodies against either anti-IGF-1R or anti-INSR verified the presence of Hybrid-R in human corneal epithelium; Hybrid-R was only activated by IGF-1. Double-labeling experiments demonstrated co-localization of IGF-1R with INSR in the nucleus. Importantly, the presence of Hybrid-R, but not IGF-1R/IGF-1R homodimer, was detected in nuclear extracts of hTCEpi cells. DNA sequencing following ChIP identified novel gene targets involved in corneal epithelial cell proliferation and survival pathways.
In addition to mediating events at the plasma membrane, IGF-1R/INSR hybrid localizes to the nucleus in proliferating corneal epithelial cells and binds to genomic targets involved in proliferation. Taken together, these data suggest a role for Hybrid-R in mediating proliferative events in the corneal epithelium.
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