期刊
AMERICAN JOURNAL OF PHYSIOLOGY-RENAL PHYSIOLOGY
卷 282, 期 5, 页码 F785-F794出版社
AMER PHYSIOLOGICAL SOC
DOI: 10.1152/ajprenal.00287.2001
关键词
endocytosis; clathrin; tubular reabsorption
资金
- NIDDK NIH HHS [DK54933, DK55389] Funding Source: Medline
Myosin VI is a reverse-direction molecular motor implicated in membrane transport events. Because myosin VI is most highly expressed in the kidney, we investigated its renal localization by using high-resolution immunocytochemical and biochemical methods. Indirect immunofluorescence microscopy revealed myosin VI at the base of the brush border in proximal tubule cells. Horseradish peroxidase uptake studies, which labeled endosomes, and double staining for clathrin adapter protein-2 showed that myosin VI was closely associated with the intermicrovillar (IMV) coated-pit region of the brush border. Localization of myosin VI to the IMV region was confirmed at the electron microscopic level by colloidal gold labeling of ultrathin cryosections. In addition, antigen retrieval demonstrated a small but significant pool of myosin VI on the microvilli. To confirm the association of myosin VI with the IMV compartment, these membranes were separated from other membrane compartments by using 15-25% OptiPrep density gradients. Immunoblotting of the gradient fractions confirmed that myosin VI was enriched with markers for the IMV microdomain of the brush border, suggesting that myosin VI associates with proteins in this compartment. Finally, we examined the expression of myosin VI during nephron development. We found myosin VI present in a diffuse cytoplasmic pattern at stage II (S-shaped body phase) and that it was only redistributed fully to the brush border in the stage IV nephron. These studies support a model for myosin VI function in the endocytic process of the proximal tubule.
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