Insulin Receptors and Downstream Substrates Associate with Membrane Microdomains after Treatment with Insulin or Chromium(III) Picolinate |
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Authors: | Abeer Al-Qatati Peter W Winter Amber L Wolf-Ringwall Pabitra B Chatterjee Alan K Van Orden Debbie C Crans Deborah A Roess B George Barisas |
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Institution: | (1) Department of Biomedical Sciences, Colorado State University, Fort Collins, CO 80523, USA;(2) Cell and Molecular Biology Program, Colorado State University, Fort Collins, CO 80523, USA;(3) Department of Chemistry, Colorado State University, Campus Mail 1872, Fort Collins, CO 80523, USA; |
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Abstract: | We have examined the association of insulin receptors (IR) and downstream signaling molecules with membrane microdomains in
rat basophilic leukemia (RBL-2H3) cells following treatment with insulin or tris(2-pyridinecarbxylato)chromium(III) (Cr(pic)3). Single-particle tracking demonstrated that individual IR on these cells exhibited reduced lateral diffusion and increased
confinement within 100 nm-scale membrane compartments after treatment with either 200 nM insulin or 10 μM Cr(pic)3. These treatments also increased the association of native IR, phosphorylated insulin receptor substrate 1 and phosphorylated
AKT with detergent-resistant membrane microdomains of characteristically high buoyancy. Confocal fluorescence microscopic
imaging of Di-4-ANEPPDHQ labeled RBL-2H3 cells also showed that plasma membrane lipid order decreased following treatment
with Cr(pic)3 but was not altered by insulin treatment. Fluorescence correlation spectroscopy demonstrated that Cr(pic)3 did not affect IR cell-surface density or compete with insulin for available binding sites. Finally, Fourier transform infrared
spectroscopy indicated that Cr(pic)3 likely associates with the lipid interface in reverse-micelle model membranes. Taken together, these results suggest that
activation of IR signaling in a cellular model system by both insulin and Cr(pic)3 involves retention of IR in specialized nanometer-scale membrane microdomains but that the insulin-like effects of Cr(pic)3 are due to changes in membrane lipid order rather than to direct interactions with IR. |
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