Klotho Coreceptors Inhibit Signaling by Paracrine Fibroblast Growth Factor 8 Subfamily Ligands

Abstract

e It has been recently established that Klotho coreceptors associate with fibroblast growth factor (FGF) receptor tyrosine kinases (FGFRs) to enable signaling by endocrine-acting FGFs. However, the molecular interactions leading to FGF-FGFR-Klotho ternary complex formation remain incompletely understood. Here, we show that in contrast to ␣Klotho, ␤Klotho binds its cognate endocrine FGF ligand (FGF19 or FGF21) and FGFR independently through two distinct binding sites. FGF19 and FGF21 use their respective C-terminal tails to bind to a common binding site on ␤Klotho. Importantly, we also show that Klotho coreceptors engage a conserved hydrophobic groove in the immunoglobulin-like domain III (D3) of the "c" splice isoform of FGFR. Intriguingly, this hydrophobic groove is also used by ligands of the paracrine-acting FGF8 subfamily for receptor binding. Based on this binding site overlap, we conclude that while Klotho coreceptors enhance binding affinity of FGFR for endocrine FGFs, they actively suppress binding of FGF8 subfamily ligands to FGFR. Fibroblast growth factor (FGF) signaling plays pleiotropic roles in metazoan development and metabolism (5, 24). Based on sequence homology and phylogenetic and structural considerations, the 18 mammalian FGF ligands are grouped into five paracrine-acting subfamilies and one endocrine-acting subfamily comprising FGF19, FGF21, and FGF23 (24,43). Paracrine FGFs have high affinity for pericellular heparan sulfate (HS) glycosaminoglycans (3, 5) and form distinct morphogenetic gradients in the pericellular matrix (27, 39) to fulfill essential roles during embryonic development (7,24,65). In contrast, endocrine FGFs exhibit poor affinity for HS (3, 15) and thus are able to enter the blood circulation to regulate key metabolic processes, including bile acid homeostasis (18,20,36) and hepatic glucose and protein metabolism (30, 57) (FGF19), glucose and lipid metabolism (4, 21, 29, 58) (FGF21), and vitamin D and phosphate homeostasis (1, 59, 62) (FGF23). These hormone-like FGFs have taken center stage in drug discovery for a number of inherited and acquired metabolic disorders (5, 6).Mammalian FGFs signal through four FGF receptor (FGFR) tyrosine kinases (FGFR1 to FGFR4) and their alternatively spliced isoforms (23, 43). The extracellular domain of a prototypical FGFR consists of three immunoglobulin-like domains (D1 to D3), and structural and biochemical studies have established that the region including D2, D3, and the D2-D3 linker comprises the minimal ligand-binding domain (54,55,63). The ligand-binding specificity of FGFR1 to FGFR3 is primarily regulated by a tissuespecific splicing in D3 of these receptors that generates "b" and "c" isoforms (9,26,42,52). Structural studies have revealed that this splicing alters the primary amino acid sequence of key ligandbinding loops/pockets in D3 (50,74).In addition to mediating paracrine FGF gradient formation in the pericellular matrix, HS is required for enhancing the affinity of paracrine FGF for FGFR and promoting dimerization of ligan...

FGF21-receptor agonists: an emerging therapeutic class for obesity-related diseases

“…mimAb1 has an ability to compete with FGF21 for KLB binding. Since FGF21 and FGF19 bind to a common site on KLB [87], mimAb1 likely interferes with endogenous FGF19's ability to bind and activate the KLB/FGFR4 complex. This may explain how mimAb1 induced weight loss without affecting adiponectin.…”

Dysregulation of phosphate metabolism and conditions associated with phosphate toxicity

“…16 The FGF23 gene has three exons separated by two introns, and the gene encodes a 32-kDa glycoprotein containing 251 amino acid residues: 24 amino acids constitute a hydrophobic signal sequence, an N terminus of 155 amino acids constitute the FGF core homology region and 72 amino acids form the C-terminal domain. [17][18][19][20] After cleavage of the 24-amino-acid signal sequence, the mature protein (25-251)-FGF23 is secreted into the circulation where three distinct forms of the FGF23 protein can be detected: a full-length mature form (25-251)-FGF23, a shorter form (25-179)-FGF23 lacking the unique C-terminal tail of 72 amino acids and a C-terminal tail. The shorter form without a C-terminal tail arises from proteolytic cleavage at the 176RXXR179 site, [17][18][19][20] and it can also be detected in the serum.…”

“…[17][18][19][20] After cleavage of the 24-amino-acid signal sequence, the mature protein (25-251)-FGF23 is secreted into the circulation where three distinct forms of the FGF23 protein can be detected: a full-length mature form (25-251)-FGF23, a shorter form (25-179)-FGF23 lacking the unique C-terminal tail of 72 amino acids and a C-terminal tail. The shorter form without a C-terminal tail arises from proteolytic cleavage at the 176RXXR179 site, [17][18][19][20] and it can also be detected in the serum. Recent studies have shown that the C terminus is not only the potential alpha-klotho-interacting site, but it also determines the functionality of the FGF23 protein.…”

“…The chimeric protein containing the N terminus of FGF2 and C terminus of FGF23 could act as a phosphatonin and could reduce the renal expression of 1a(OH)ase. 19,20 It is worth mentioning that the presence of the C terminus of FGF23 in the chimeric protein paved the way for the alpha-klotho interaction. Recently, a family with sequence similarity 20, member C (Fam20C), was shown to phosphorylate FGF23.…”

Novel strategies and therapeutic options for the management of primary biliary cholangitis

“…It works by altering the activity of the CYP7A1 enzyme through binding to the FGFR4-β-klotho co-receptor complex in the liver, release of which is promoted by FXR activity, and hence, essentially acting in a similar fashion to OCA but at a slightly different target point in the pathway. 58 This interruption in the enterohepatic circulation pathway and resulting changes in the FGF-19 activity leads to significant alterations in both the size and composition of the BA pool. NGM-282 effectively mimics the actions of FGF-19 on BA synthesis through the binding of FGFR4c-β-klotho co-receptor.…”