Mitotic CDK1 and 4E-BP1 II: A single phosphomimetic mutation in 4E-BP1 induces glucose intolerance in mice

OBJECTIVE: Cyclin-dependent kinase 1 (CDK1)/cyclin B1 phosphorylates many of the same substrates as mTORC1 (a key regulator of glucose metabolism), including the eukaryotic initiation factor 4E-binding protein 1 (4E-BP1). Only mitotic CDK1 phosphorylates 4E-BP1 at residue S82 in mice (S83 in humans), in addition to the common 4E-BP1 phospho-acceptor sites phosphorylated by both CDK1 and mTORC1. We examined glucose metabolism in mice having a single aspartate phosphomimetic amino acid knock in substitution at the 4E-BP1 serine 82 (4E-BP1(S82D)) mimicking constitutive CDK1 phosphorylation. METHODS: Knock-in homozygous 4E-BP1(S82D) and 4E-BP1(S82A) C57Bl/6N mice were assessed for glucose tolerance testing (GTT) and metabolic cage analysis on regular and on high-fat chow diets. Gastrocnemius tissues from 4E-BP1(S82D) and WT mice were subject to Reverse Phase Protein Array analysis. Since the bone marrow is one of the few tissues typically having cycling cells that transit mitosis, reciprocal bone-marrow transplants were performed between male 4E-BP1(S82D) and WT mice, followed by metabolic assessment, to determine the role of actively cycling cells on glucose homeostasis. RESULTS: Homozygous knock-in 4E-BP1(S82D) mice showed glucose intolerance that was markedly accentuated with a diabetogenic high-fat diet (p = 0.004). In contrast, homozygous mice with the unphosphorylatable alanine substitution (4E-BP1(S82A)) had normal glucose tolerance. Protein profiling of lean muscle tissues, largely arrested in G(0), did not show protein expression or signaling changes that could account for these results. Reciprocal bone-marrow transplantation between 4E-BP1(S82D) and wild-type littermates revealed a trend for wild-type mice with 4E-BP1(S82D) marrow engraftment on high-fat diets to become hyperglycemic after glucose challenge. CONCLUSIONS: 4E-BP1(S82D) is a single amino acid substitution that induces glucose intolerance in mice. These findings indicate that glucose metabolism may be regulated by CDK1 4E-BP1 phosphorylation independent from mTOR and point towards an unexpected role for cycling cells that transit mitosis in diabetic glucose control.