An optimized fractionation method reveals insulin-induced membrane surface localization of GLUT1 to increase glycolysis in LβT2 cells

Mol Cell Endocrinol. 2025 Jan 1:595:112405. doi: 10.1016/j.mce.2024.112405. Epub 2024 Oct 29.

Abstract

Insulin is an important regulator of whole-body glucose homeostasis. In insulin sensitive tissues such as muscle and adipose, insulin induces the translocation of glucose transporter 4 (GLUT4) to the cell membrane, thereby increasing glucose uptake. However, insulin also signals in tissues that are not generally associated with glucose homeostasis. In the human reproductive endocrine axis, hyperinsulinemia suppresses the secretion of gonadotropins from gonadotrope cells of the anterior pituitary, thereby linking insulin dysregulation to suboptimal reproductive health. In the mouse, gonadotropes express the insulin receptor which has the canonical signaling response of IRS, AKT, and mTOR activation. However, the functional outcomes of insulin action on gonadotropes are unclear. Here, we demonstrate through use of an optimized cell fractionation protocol that insulin stimulation of the LβT2 gonadotropic cell line results in the unexpected translocation of GLUT1 to the plasma membrane. Using our high purity fractionation protocol, we further demonstrate that though Akt signaling in response to insulin is intact, insulin-induced translocation of GLUT1 occurs independently of Akt activation in LβT2 cells.

Keywords: Cytosol; Endosomes; Glucose transporter; Gonadotrope; Insulin; Membrane; Nuclear; Phosphorylated akt; Subcellular location.

MeSH terms

  • Animals
  • Cell Fractionation / methods
  • Cell Line
  • Cell Membrane* / metabolism
  • Glucose / metabolism
  • Glucose / pharmacology
  • Glucose Transporter Type 1* / metabolism
  • Glycolysis* / drug effects
  • Insulin* / metabolism
  • Insulin* / pharmacology
  • Mice
  • Protein Transport / drug effects
  • Proto-Oncogene Proteins c-akt* / metabolism
  • Signal Transduction / drug effects

Substances

  • Insulin
  • Glucose Transporter Type 1
  • Proto-Oncogene Proteins c-akt
  • Glucose