Tyrosine sulfation is required for agonist recognition by glycoprotein hormone receptors

Abstract

The glycoprotein hormone receptors (thyrotrophin receptor, TSHr; luteinizing hormone/chorionic gonadotrophin receptor, LH/CGr; follicle-stimulating hormone receptor, FSHr) constitute a subfamily of rhodopsin-like G protein-coupled receptors (GPCRs) with a long N-terminal extracellular extension responsible for high-af®nity hormone binding. These ectodomains contain two cysteine clusters¯anking nine leucine-rich repeats (LRR), a motif found in several protein families involved in protein±protein interactions. Similar to the situation described recently in CCR5, we demonstrate here that the TSHr, as it is present at the cell surface, is sulfated on tyrosines in a motif located downstream of the C-terminal cysteine cluster. Sulfation of one of the two tyrosines in the motif is mandatory for high-af®nity binding of TSH and activation of the receptor. Site-directed mutagenesis experiments indicate that the motif, which is conserved in all members of the glycoprotein hormone receptor family, seems to play a similar role in the LH/CG and FSH receptors.

Structure of follicle-stimulating hormone in complex with the entire ectodomain of its receptor

“…The large number of hydrophobic residues creates a low dielectric-constant microenvironment, enhancing Coulomb charge-charge attraction between the FSHR sulfate and FSH R35β aligned on the upper side of the binding pocket, as well as providing a necessary milieu to accommodate the Y335's hydrophobic phenyl ring. Despite the small interaction area at this site, sulfated Y335 is essential for FSHR activation (16,23). In LHR, the equivalently sulfated tyrosine Y331 and its preceding residue D330 have also been determined as the key signaling residues by site-directed mutagenesis (22).…”

“…In the absence of this domain, it is difficult to understand how the ligand-binding signal is conveyed to the transmembrane domain, and the observed ligand receptorbinding mode was challenged and an alternative mode was proposed (15). It has also been shown that sulfated tyrosines in the hinge region of GPHRs (including Y335 for FSHR) are required for hormone recognition and signaling (16). How sulfated residues interact with the hormones, and subsequently lead to receptor activation, remains poorly understood.…”

Tyrosine sulfation in N-terminal domain of human C5a receptor is necessary for binding of chemotaxis inhibitory protein of Staphylococcus aureus

“…This modification tends to occur in acidic regions of proteins, usually containing multiple tyrosines [9,11] . Several receptors for chemokines and hormones, including CCR5, CXCR3, C3XCR1, and thyroid-stimulating hormone receptor, have been shown to be sulfated on tyrosines in their amino terminal extracellular domains and some of this sulfation is critical for ligand binding [12][13][14][15][16] . Amino-terminal sulfation of tyrosines in the chemotactic receptor for C5a also contributes to formation of the docking site for the C5a anaphylatoxin [17] .…”

Female Infertility in grt Mice is Caused by Thyroid Hormone Deficiency, not by Insufficient TPST2 Activity in the Reproductive Organs

“…Although TPST1 and 2 in mammals have 65~67% identical amino acid sequences [28], they seem to have different acceptor preferences [31,37]. Evidence indicates that the posttranslational modification by tyrosine sulfation regulates many important protein-protein interactions and modulates binding affinity and specificity [8,10,11,18,24,26,29,[33][34][35][36]. We previously showed that the sulfation of the tyrosine 385 of TSHR by TPST2 is indispensable for the activation of TSH signaling, and grt mice develop hypothyroidism and dwarfism since they are unable to fully respond to TSH.…”

“…The expression of TPST2 in the reproductive organs raises the possibility of TPST2 having local functions in mice. Interestingly, luteinizing hormone receptor (LHR) and follicle-stimulating hormone receptor (FSHR), which are essential to the female reproductive systems, have been recently reported to be tyrosine-sulfated [4,36] and the tyrosine-sulfated products have been predicted to modify agonist recognition [10]. Several reports have indicated that the deficiency of LHR or FSHR signal cascades causes female infertility, which is characterized by follicular arrest beyond the Graafian follicle and disturbed ovulation [1,23,38,42].…”