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Na(+)-coupled alanine transport in LLC-PK1 cells
G A Kimmich1, J Randles, J Wilson
1Department of Biochemistry, School of Medicine and Dentistry, University of Rochester, New York 14642.
The American Journal of Physiology
|October 1, 1994
Summary
This study reveals that alanine transport in renal cells shifts to the ASC system as cells mature. This transport is sodium-dependent and differs significantly from glucose transporter mechanisms.
Area of Science:
- Cell Biology
- Renal Physiology
- Molecular Transport
Background:
- Alanine (Ala) transport is crucial in renal epithelial cells.
- Cellular differentiation, specifically postconfluence, alters nutrient transport mechanisms.
Purpose of the Study:
- To characterize alanine transport kinetics and identify the specific transporter system in LLC-PK1 cells.
- To compare alanine transport with alpha-methylglucoside (AMG) transport under varying conditions.
Main Methods:
- Utilized ATP-depleted LLC-PK1 cells to control membrane potential and Na+ gradients.
- Performed kinetic analysis of alanine and AMG influx with varying substrate and Na+ concentrations.
- Investigated substrate inhibition patterns and the effect of membrane potential on transport.
Main Results:
- Alanine transport decreased by 75% in postconfluent cells, while Na(+)-coupled AMG transport increased fourfold.
- Alanine influx is primarily Na+-dependent, with low affinity for Li+, and exhibits characteristics of the ASC amino acid transporter.
- Kinetic analysis revealed a Michaelis constant (Km) for alanine of 380 µM and for Na+ of 32 mM, with a transport stoichiometry of 1:1.
- AMG transport showed a sigmoidal relationship with Na+ concentration and a stoichiometry of 2:1.
Conclusions:
- Alanine transport in LLC-PK1 cells transitions to the ASC system in postconfluent cultures.
- Differences in Na+ coupling stoichiometry (1:1 for Ala, 2:1 for AMG) explain variations in transport system interactions.