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KMID : 0894520170210020111
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Development & Reproduction
2017 Volume.21 No. 2 p.111 ~ p.120
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Internalization of Rat FSH and LH/CG Receptors by rec-eCG in CHO-K1 Cells
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Park Jong-Ju
Seong Hun-Ki
Kim Jeong-Soo
Munkhzaya Byambaragchaa
Kang Myung-Hwa
Min Kwan-Sik
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Abstract
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Equine chorionic gonadotropin (eCG) is a unique molecule that elicits the response characteristics of both fol-licle-stimulating hormone (FSH) and luteinizing hormone (LH) in other species. Previous studies from this laboratory had demonstrated that recombinant eCG (rec-eCG) from Chinese hamster ovary (CHO-K1) cells exhibited both FSH- and LH-like activity in rat granulosa and Leydig cells. In this study, we analyzed receptor internalization through rec-eCGs, wild type eCG (eCGβ/α) and mutant eCG (eCGβ/αΔ56) with an N-linked oligosaccharide at Asn56 of the α-subunit. Both the rec-eCGs were obtained from CHO-K1 cells. The agonist activation of receptors was analyzed by measuring stimulation time and concentra-tions of rec-eCGs. Internalization values in the stably selected rat follicle-stimulating hormone receptor (rFSHR) and rat lute-inizing/chorionic gonadotropin receptor (rLH/CGR) were highest at 50 min after stimulation with 10 ng of rec-eCGβ/α. The dose-dependent response was highest when 10 ng of rec-eCGβ/α was used. The deglycosylated eCGβ/αΔ56 mutant did not enhance the agonist-stimulated internalization. We concluded that the state of activation of rFSHR and rLH/CGR could be modulated through agonist-stimulated internalization. Our results suggested that the eLH/CGRs are mostly internalized within 60 min by agonist-stimulation by rec-eCG. We also suggested that the lack of responsiveness of the deglycosylated eCGβ/ αΔ56 was likely because the site of glycosylation played a pivotal role in agonist-stimulated internalization in cells express-ing rFSHR and rLH/CGR.
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KEYWORD
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rec-eCGβ/α, Deglycosylated mutants, Internalization, rFSHR, rLH/CGR
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