FGF21 is an endocrine signal of protein restriction

Abstract

Enhanced fibroblast growth factor 21 (FGF21) production and circulation has been linked to the metabolic adaptation to starvation. Here, we demonstrated that hepatic FGF21 expression is induced by dietary protein restriction, but not energy restriction. Circulating FGF21 was increased 10-fold in mice and rats fed a low-protein (LP) diet. In these animals, liver Fgf21 expression was increased within 24 hours of reduced protein intake. In humans, circulating FGF21 levels increased dramatically following 28 days on a LP diet. LP-induced increases in FGF21 were associated with increased phosphorylation of eukaryotic initiation factor 2α (eIF2α) in the liver, and both baseline and LP-induced serum FGF21 levels were reduced in mice lacking the eIF2α kinase general control nonderepressible 2 (GCN2). Finally, while protein restriction altered food intake, energy expenditure, and body weight gain in WT mice, FGF21-deficient animals did not exhibit these changes in response to a LP diet. These and other data demonstrate that reduced protein intake underlies the increase in circulating FGF21 in response to starvation and a ketogenic diet and that FGF21 is required for behavioral and metabolic responses to protein restriction. FGF21 therefore represents an endocrine signal of protein restriction, which acts to coordinate metabolism and growth during periods of reduced protein intake.

Figure 6. Hypothetical model of protein-dependent regulation of FGF21 and its effects on food intake and EE.

Reduced consumption of dietary protein leads to reduced delivery of amino acids to the liver, activating the kinase GCN2 and leading to increased eIF2α phosphorylation and activation of ATF4/ATF5. ATF4/5 bind AAREs within the FGF21 promoter, leading to increased liver FGF21 production and increased circulating FGF21. PPARα is also required for normal levels of circulating FGF21. Finally, increased circulating levels of FGF21 increase EE and food intake and also alter body weight gain and body composition.

Figure 5. FGF21 is required for LP-induced changes in EE, food intake, body weight, and body composition.

EE in WT (A) and Fgf21-KO (B) mice consuming control or LP diet. Average EE (C) and change in EE from baseline (D) over days 5 to 7 of diet consumption. Average daily food intake (E) in WT and Fgf21-KO mice over the 14 days of diet exposure. Change in body weight (F), fat mass (G), and lean mass (H) in WT and Fgf21-KO mice on control or LP diet for 14 days. n = 10/group, *P < 0.05; **P < 0.01.

Figure 4. LP-induced increases in circulating FGF21 depend on both GCN2 and PPARα signaling.

Liver Ppara mRNA expression was assessed in mice and rats on LP diet for 14 days (A) as well as liver expression of PPARα responsive genes in rats (B). Western blot of eIF2α phosphorylation in liver of rats consuming control or LP diet for 2 days (C), and eIF2α phosphorylation in liver of mice and rats during various models of protein restriction described previously (D). Serum FGF21 (E) and liver eIF2α phosphorylation (F) was measured in WT, Gcn2-KO, and Ppara-KO mice following 4 days of LP diet. n = 5–8/group. *P < 0.05; **P < 0.01 vs. control; ^#P < 0.05 vs. WT control.

Figure 3. Dietary protein restriction increases FGF21 in humans.

Plasma FGF21 concentrations were measured in human subjects given isocaloric amounts of diets that were either normal (A) or low (B) in protein for 28 days, and the change in FGF21 between baseline (day 0) and day 28 was compared (C). n = 8–9/group; *P = 0.01.

Figure 2. Protein restriction underlies the increase in FGF21 during food restriction or a KD.

Serum FGF21 (A) and liver Fgf21 mRNA expression (B) were compared in rats placed on diets that independently restricted energy vs. protein for 4 days: control (NP:NE), LP:NE, NP:LE, and LP:LE. Serum FGF21 was also compared in a separate group of rats (C) that were food deprived for 48 hours and then refed equal caloric amounts of either a LP or HP diet. Serum BHBA (D) and FGF21 (E) were measured in mice fed control, KD, or KD supplemented with an isocaloric amount of either carbohydrate (KD + CHO) or protein (KD + P) for 4 days. n = 5/group; *P < 0.05; **P < 0.01; ***P < 0.001 vs. control/NP:NE; ^#P < 0.05 vs. KD.

Figure 1. FGF21 is increased by dietary protein restriction in rodents.

Rats (A) and mice (B) were placed on isocaloric control or LP diets for 4 or 14 days, with circulating FGF21 protein levels assessed by ELISA. Fgf21 mRNA expression was measured in rat liver, eWAT, and muscle (C) following 4 days of LP. Acute changes in liver Fgf21 mRNA expression in rats were measured over time (D). n = 5–8/group; *P < 0.05; **P < 0.01; ***P < 0.001 vs. control.