Understanding the Physical Interactions in the FGF21/FGFR/β‐Klotho Complex: Structural Requirements and Implications in FGF21 Signaling

Abstract

The endocrine fibroblast growth factor 21 (FGF21) requires both fibroblast growth factor receptor (FGFR) and b-Klotho for signaling. In this study, we sought to understand the inter-molecular physical interactions in the FGF21 ⁄ FGFR ⁄ b-Klotho complex by deleting key regions in FGFR1c or FGF21. Deletion of the D1 and the D1-D2 linker (the D1 ⁄ linker region) from FGFR1c led to b-Klotho-independent receptor activation by FGF21, suggesting that there may be a direct interaction between FGF21 and the D1 ⁄ linker region-deficient FGFR1c. Consistent with this, the extracellular portion of FGFR1c lacking the D1 ⁄ linker region blocked FGF21 action in a reporter assay, presumably by binding to and sequestering FGF21 from acting on cell surface receptor complex. In addition, the D1 ⁄ linker region-deficient FGFR1c had enhanced interaction with b-Klotho. Further, we demonstrated that deletion of the D1 ⁄ linker region enhanced the formation of the FGF21 ⁄ b-Klotho ⁄ FGFR1c ternary complex in both Biacore and asymmetrical flow field flow fraction-ation studies. Finally, we found that the N-terminus of FGF21 is involved in the interaction with FGFR1c and FGF21 ⁄ b-Klotho ⁄ FGFR1c ternary complex formation. Taken together, our data suggest that the D1 ⁄ linker region regulates both the FGF21 ⁄ FGFR1c and FGFR1c ⁄ b-Klotho interaction, and a direct interaction of FGF21 with FGFR1c may be an important step in receptor-mediated FGF21 signaling. Key words: D1 and the D1-D2 linker, FGF21, FGFR1c, b-Klotho Abbreviations: AF4, asymmetrical flow field flow fractionation; D1, immunoglobin domain 1; FGF, fibroblast growth factor; FGFR, fibroblast growth factor receptor; HSPG, heparin sulfate proteoglycan; mIMCD3, murine renal inner medullary collecting duct; the D1 ⁄ linker region, D1 and the D1-D2 linker. Fibroblast growth factor 21 (FGF21) and its two other subfamily members, FGF19 and FGF23, have emerged as potent endocrine hormones involved in the regulation of diverse physiological functions (1-3). In particular, FGF21 has generated a lot of excitement for its potential of becoming a therapeutic for type 2 diabetes and dyslipi-demia (4). Noticeably, administration of recombinant FGF21 lowered plasma glucose, insulin, triglycerides, and cholesterol levels and improved insulin sensitivity, hepatic steatosis, and obesity in a range of diabetes ⁄ obesity animal models (3,5-7). The key differences that distinguish the FGF19 subfamily members from paracrine FGFs are their lack of or weak heparin-binding properties (8) and their dependence on Klotho family transmembrane proteins as co-receptor for the activation of FGF receptor (FGFR) sig-naling (9). a-Klotho is required for FGF23 to regulate renal phosphate metabolism (10), whereas b-Klotho is required for FGF21 to regulate glucose and lipid metabolism (11-14) and FGF19 to regulate bile acid metabolism (15,16). Interestingly, FGF19 can also induce signaling with a-Klotho as a co-receptor, (16) although the physiological implication is still unclear. In contrast, paracrine FGFs signal...

Unliganded Fibroblast Growth Factor Receptor 1 Forms Density-independent Dimers

“…Previously, co-immunoprecipitation was used to demonstrate the ligand-independent formation of stable FGFR1c⅐KLB complex (9,10,58). However, a model has been recently proposed in which FGF21 would facilitate FGFR1c⅐KLB interaction by suppressing intramolecular interactions between FGFR1c D1 and the rest of the ECD (28). Stabilization of the interaction between FGFR1c and KLB by FGF21 was also demonstrated utilizing recombinant protein fragments (28).…”

“…For example, FGFR1 and KLB can be co-immunoprecipitated, suggesting a formation of ligand-independent complex (9, 10). However, surface plasmon resonance sensorgrams showed that interaction between FGFR ECD and KLB ECD is very weak unless stabilized by FGF21 (28).…”

The Future of Genomic Medicine Involves the Maintenance of Sirtuin 1 in Global Populations

“…38 FGF21 binds to FGF receptor and beta klotho receptor complex [38][39][40][41][42][43] and activates adipose tissue Sirt1 by increases in NAD+ and activation ofPGC1-alpha and AMP activated protein kinase (AMPK). 37,44 FGF21 and its effect on thermoregulation 45 may involve Sirt 1 regulation of heat shock proteins (HSP) by deacetylation of heat shock factor (HSF) via PGC1α as a critical repressor of HSF1-mediated transcriptional programs.…”

TCF4/β‑catenin complex is directly upstream of FGF21 in mouse stomach cancer cells

“…FGFs have been demonstrated to be associated with the processes of embryogenesis, gastrulation, somitogenesis, body plan formation, organogenesis and skin wound healing (2)(3)(4)(5)(6)(7). FGF21 is a member of the FGF family that belongs to the FGF19 subfamily, and activation of the FGF receptor by FGF21 requires the co-receptor β-klotho (8,9). FGF21 has been reported to preferentially express in the liver (10).…”