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Receptor-mediated vectorial transcytosis of epidermal growth factor by Madin-Darby canine kidney cells
E Maratos-Flier1, C Y Kao, E M Verdin
1Elliott P. Joslin Research Laboratory, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts 02215.
Abstract:
Transcellular transport of a variety of ligands may be an important mechanism by which regulatory substances reach their site of action. We have studied the transcellular transport of two 6,000-mol-wt proteins, epidermal growth factor (EGF) and insulin, across polarized Madin-Darby canine kidney (MDCK) cells grown on dual-sided chambers on a nitrocellulose filter substrate. When grown on these chambers, MDCK cells are polarized and express distinct basal and apical surfaces. MDCK cells are capable of unidirectional transport of EGF from the basal-to-apical direction, 50% of bound EGF transported in 2 h. Transport was inhibited by the addition of unlabeled EGF in a dose-dependent manner. Anti-EGF receptor Ab, which inhibited binding, also inhibited transport. No transport in the apical-to-basal direction is noted. Insulin transport is not observed in either direction. Transport correlates with the presence of ligand-specific receptors on the cell surface. Hence, EGF receptors (Ro = 48,000, Kd = 3.5 X 10(-10) M) are found only on the basal surface of the MDCK cells and neither surface expresses insulin receptors. Characterization of the EGF receptors on MDCK cells, as assessed by affinity, molecular mass, and anti-receptor antibody binding reveals that this receptor has similar characteristics to EGF receptors previously described on a variety of cells. Hence, the EGF receptor can function as a transporter of EGF across an epithelial cell barrier.
Insights
Epidermal growth factor (EGF) is transported across kidney cells via its receptor, but insulin is not. This demonstrates how specific receptors facilitate transcellular transport of regulatory molecules.
Area of Science:
- Cell Biology
- Molecular Biology
- Physiology
Background:
- Transcellular transport is crucial for distributing regulatory substances.
- Polarized epithelial cells present distinct apical and basal surfaces.
- Understanding ligand transport mechanisms is key to cellular communication.
Purpose of the Study:
- To investigate the transcellular transport of epidermal growth factor (EGF) and insulin across polarized Madin-Darby canine kidney (MDCK) cells.
- To determine the role of ligand-specific receptors in this transport process.
Main Methods:
- Utilized polarized MDCK cells grown on dual-sided chambers with a nitrocellulose filter substrate.
- Assessed unidirectional transport of EGF and insulin across cell layers.
- Investigated the effect of unlabeled EGF and anti-EGF receptor antibodies on EGF transport.
- Characterized EGF receptor presence and properties on MDCK cell surfaces.
Main Results:
- MDCK cells exhibited unidirectional transport of EGF from the basal to apical surface (50% in 2 hours).
- EGF transport was dose-dependently inhibited by unlabeled EGF and anti-EGF receptor antibodies.
- No significant transport of insulin was observed in either direction.
- EGF receptors were localized exclusively to the basal surface of MDCK cells; insulin receptors were absent.
Conclusions:
- The EGF receptor functions as a transporter for EGF across an epithelial cell barrier.
- Transcellular transport of ligands like EGF is mediated by specific cell surface receptors.
- The polarity of receptor expression dictates the directionality of ligand transport.