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Published on: November 16, 2011
Mitotic CDK1 and 4E-BP1 II: A single phosphomimetic mutation in 4E-BP1 induces glucose intolerance in mice
Simon Cao1,2, Michael J Jurczak3,4, Yoko Shuda1
1Hillman Cancer Center, Cancer Virology Program, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
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-BP1S82D) mimicking constitutive CDK1 phosphorylation.
Methods:
Knock-in homozygous 4E-BP1S82D and 4E-BP1S82A 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-BP1S82D 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-BP1S82D and WT mice, followed by metabolic assessment, to determine the role of actively cycling cells on glucose homeostasis.
Results:
Homozygous knock-in 4E-BP1S82D 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-BP1S82A) had normal glucose tolerance. Protein profiling of lean muscle tissues, largely arrested in G0, did not show protein expression or signaling changes that could account for these results. Reciprocal bone-marrow transplantation between 4E-BP1S82D and wild-type littermates revealed a trend for wild-type mice with 4E-BP1S82D marrow engraftment on high-fat diets to become hyperglycemic after glucose challenge.
Conclusions:
4E-BP1S82D 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.
Insights
Mice with a specific 4E-BP1 modification mimicking CDK1 phosphorylation showed glucose intolerance, especially on high-fat diets. This suggests CDK1-mediated 4E-BP1 phosphorylation regulates glucose metabolism, potentially involving cycling cells.
Area of Science:
- Cellular Biology
- Metabolic Regulation
- Molecular Signaling
Background:
- Cyclin-dependent kinase 1 (CDK1)/cyclin B1 and mTORC1 regulate glucose metabolism by phosphorylating shared substrates like 4E-binding protein 1 (4E-BP1).
- CDK1 uniquely phosphorylates 4E-BP1 at serine 82 (S82) during mitosis, distinct from mTORC1 phosphorylation sites.
Purpose of the Study:
- To investigate the role of CDK1-mediated 4E-BP1 phosphorylation on glucose metabolism.
- To examine the impact of a phosphomimetic substitution at 4E-BP1 S82 (4E-BP1S82D) on glucose homeostasis in mice.
Main Methods:
- Generated knock-in mice with 4E-BP1 S82 phosphomimetic (4E-BP1S82D) or unphosphorylatable (4E-BP1S82A) substitutions.
- Conducted glucose tolerance tests (GTT) and metabolic cage analyses on standard and high-fat diets.
- Performed reciprocal bone marrow transplants between 4E-BP1S82D and wild-type mice to assess the role of cycling cells.
Main Results:
- Homozygous 4E-BP1S82D mice exhibited glucose intolerance, significantly worsened by a high-fat diet (p = 0.004).
- Conversely, 4E-BP1S82A mice displayed normal glucose tolerance.
- Bone marrow transplantation studies indicated a trend towards hyperglycemia in wild-type mice receiving 4E-BP1S82D marrow on high-fat diets.
Conclusions:
- Constitutive CDK1 phosphorylation at 4E-BP1 S82 induces glucose intolerance in mice, independent of mTORC1.
- These findings highlight a novel role for CDK1-mediated 4E-BP1 phosphorylation in glucose metabolism and suggest an involvement of mitotic cycling cells in diabetic glucose control.
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