Our long-term goal is to determine how gonadotropins function. Gonadotropins require N-glycans to fully activate their cognate receptors even though high affinity binding involves only protein-protein interactions. Gonadotropins are used to treat infertility, but become progressively less effective as patients age. Urinary and recombinant hFSH preparations lack a novel, highly active glycoform, di-glycosylated hFSH, that possesses only a subunit N-glycans and which predominates in pituitaries of young women. The relative abundance of di-glycosylated and tetra-glycosylated hFSH appear to be physiologically regulated. This proposal will define these changes and study possible mechanisms to explain differences in activity via 2 specific aims. 1. Characterize cyclic- and age-related changes in hFSH glycoform abundance. First, we will measure urinary FSH glycoform abundance in young healthy cycling women. Our working hypothesis is that ovarian feedback will alter FSH glycoform abundance. Second, we will evaluate glycoform abundance during the perimenopausal period, as the timing of the shift in glycoform abundance is unknown. One hypothesis is that FSH glycoform abundance changes gradually during the perimenopausal period, thereby contributing to declining fertility. An alternative hypothesis is that glycoform abundance changes abruptly at the menopausal transition. Finally, we will examine glycoform abundance in post-menopausal women and compare individuals receiving estrogen replacement therapy with those receiving none. Our working hypothesis is estrogen selectively inhibits hFSHfi glycosylation. 2. Identify a and /? subunit glycosylation patterns modulating FSH receptor-binding and FSH-stimulated steroidogenesis. We will compare receptor-binding and steroidogenic activities of di- and tetra-glycosylated hFSH. Our hypothesis is that di-glycosylated hFSH on rate is faster than tetra-glycosylated hFSH. The relationship of FSHR binding and steroidogenic activity will be compared. Our working hypothesis is that di-glycosylated hFSH will exhibit greater biological activity than predicted by its significatly higher receptor-binding activity. This project is centered around glycoforms readily identified by western blotting and lays the ground for future studies by contributing to the development of methods to rapidly characterize FSH glycosylation. A perimenopausal marker and more effective preparations for treating infertility are potential practical outcomes of this project.

Agency
National Institute of Health (NIH)
Institute
National Institute on Aging (NIA)
Type
Research Program Projects (P01)
Project #
5P01AG029531-04
Application #
8377839
Study Section
Special Emphasis Panel (ZAG1-ZIJ-5)
Project Start
Project End
Budget Start
2012-04-01
Budget End
2013-03-31
Support Year
4
Fiscal Year
2012
Total Cost
$258,002
Indirect Cost
$73,895
Name
Wichita State University
Department
Type
DUNS #
053078127
City
Wichita
State
KS
Country
United States
Zip Code
67260
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Wang, Huizhen; Hastings, Richard; Miller, William L et al. (2016) Fshb-iCre mice are efficient and specific Cre deleters for the gonadotrope lineage. Mol Cell Endocrinol 419:124-38
Wang, Huizhen; May, Jacob; Butnev, Viktor et al. (2016) Evaluation of in vivo bioactivities of recombinant hypo- (FSH(21/18)) and fully- (FSH(24)) glycosylated human FSH glycoforms in Fshb null mice. Mol Cell Endocrinol 437:224-236
Wang, Huizhen; Graham, Ian; Hastings, Richard et al. (2015) Gonadotrope-specific deletion of Dicer results in severely suppressed gonadotropins and fertility defects. J Biol Chem 290:2699-714
He, C; Lv, X; Hua, G et al. (2015) YAP forms autocrine loops with the ERBB pathway to regulate ovarian cancer initiation and progression. Oncogene 34:6040-54
He, Chunbo; Mao, Dagan; Hua, Guohua et al. (2015) The Hippo/YAP pathway interacts with EGFR signaling and HPV oncoproteins to regulate cervical cancer progression. EMBO Mol Med 7:1426-49
Butnev, Viktor Y; Butnev, Vladimir Y; May, Jeffrey V et al. (2015) Production, purification, and characterization of recombinant hFSH glycoforms for functional studies. Mol Cell Endocrinol 405:42-51
Jiang, Chao; Hou, Xiaoying; Wang, Cheng et al. (2015) Hypoglycosylated hFSH Has Greater Bioactivity Than Fully Glycosylated Recombinant hFSH in Human Granulosa Cells. J Clin Endocrinol Metab 100:E852-60
Bousfield, George R; Butnev, Vladimir Y; White, William K et al. (2015) Comparison of Follicle-Stimulating Hormone Glycosylation Microheterogenity by Quantitative Negative Mode Nano-Electrospray Mass Spectrometry of Peptide-N Glycanase-Released Oligosaccharides. J Glycomics Lipidomics 5:

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