Transforming growth factor-β1 stimulates vascular smooth muscle cell l-proline transport by inducing system A amino acid transporter 2 (SAT2) gene expression

Abstract

Transforming growth factor-β1 (TGF-β1) is a multifunctional cytokine that contributes to arterial remodelling by stimulating vascular smooth muscle cell (SMC) growth and collagen synthesis at sites of vascular injury. Since l-proline is essential for the synthesis of collagen, we examined whether TGF-β1 regulates the transcellular transport of l-proline by vascular SMCs. l-Proline uptake by vascular SMCs was primarily sodium-dependent, pH-sensitive, blocked by neutral amino acids and α-(methylamino)isobutyric acid, and exhibited trans-inhibition. Treatment of SMCs with TGF-β1 stimulated l-proline transport in a concentration- and time-dependent manner. The TGF-β1-mediated l-proline uptake was inhibited by cycloheximide or actinomycin D. Kinetic studies indicated that TGF-β1-induced l-proline transport was mediated by an increase in transport capacity independent of any changes in the affinity for l-proline. TGF-β1 stimulated the expression of system A amino acid transporter 2 (SAT2) mRNA in a time-dependent fashion that paralleled the increase in l-proline transport. Reverse transcriptase PCR failed to detect the presence of SAT1 or amino acid transporter 3 (ATA3) in either untreated or TGF-β1-treated SMCs. These results demonstrate that l-proline transport by vascular SMCs is mediated predominantly by the SAT and that TGF-β1 stimulates SMC l-proline uptake by inducing the expression of the SAT2 gene. The ability of TGF-β1 to induce SAT2 expression may function to provide SMCs with the necessary levels of l-proline required for collagen synthesis and cell growth.