Dynamic tracking and mobility analysis of single GLUT4 storage vesicle in live 3T3-L1 cells.
Glucose transporter 4 (GLUT4) is responsible for insulin-stimulated glucose transporting into the insulin-sensitive fat and muscle cells. The dynamics of GLUT4 storage vesicles (GSVs) remains to be explored and it is unclear how GSVs are arranged based on their mobility. We examined this issue in 3T3-L1 cells via investigating the three-dimensional mobility of single GSV labeled with EGFP-fused GLUT4. A thin layer of cytosol right adjacent to the plasma membrane was illuminated and successively imaged at 5 Hz under a total internal reflection fluorescence microscope with a penetration depth of 136 nm. Employing single particle tracking, the three-dimensional subpixel displacement of single GSV was tracked at a spatial precision of 22 nm. Both the mean square displacement and the diffusion coefficient were calculated for each vesicle. Tracking results revealed that vesicles moved as if restricted within a cage that has a mean radius of 160 nm, suggesting the presence of some intracellular tethering matrix. By constructing the histogram of the diffusion coefficients of GSVs, we observed a smooth distribution instead of the existence of distinct groups. The result indicates that GSVs are dynamically retained in a continuous and wide range of mobility rather than into separate classes.
Li
Chen Hong
CH
Institute of Biophysics and Biochemistry, Huazhong University of Science and Technology, Wuhan 43007, China.
Bai
Li
L
Li
Dong Dong
DD
Xia
Sheng
S
Xu
Tao
T
D041721:3T3-L1 Cells; D000818:Animals; D022162:Cytoplasmic Vesicles / Q000378:metabolism*; D004058:Diffusion; D051275:Glucose Transporter Type 4; D049452:Green Fluorescent Proteins / Q000235:genetics; D051379:Mice; D018613:Microscopy, Confocal / Q000379:methods; D009004:Monosaccharide Transport Proteins / Q000235:genetics / Q000378:metabolism*; D009068:Movement; D009124:Muscle Proteins / Q000235:genetics / Q000378:metabolism*; D016011:Normal Distribution; D010316:Particle Size; D021381:Protein Transport / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D013050:Spectrometry, Fluorescence / Q000379:methods; D013997:Time Factors
D051275:Glucose Transporter Type 4; D009004:Monosaccharide Transport Proteins; D009124:Muscle Proteins; D011993:Recombinant Fusion Proteins; C495693:Slc2a4 protein, mouse; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins