Matrix Gla protein (MGP) is a mineral binding extra-cellular matrix protein synthesized by vascular smooth muscle cells (VSMCs) and chondrocytes which is thought to be a key regulator of tissue calcification. In this study we identified 4 polymorphisms in the promoter region of the human MGP gene. Transfection studies showed that the G-7A and T-138C polymorphisms have an important impact on in vitro promoter activity when transiently transfected into VSMCs. We found that one of these polymorphisms (T-138C) is significantly correlated with serum MGP levels in human subjects. Promoter deletion analysis showed that this polymorphism lies in a region of the promoter critical for transcription in VSMCs. This region contains a potential activating protein-1 (AP-1) binding element located between -142 and –136. We have demonstrated that the T-138C polymorphism results in altered binding of an AP-1 complex to this region. The –138T allelic variant binds AP-1 complexes consisting primarily of c-Jun, JunB and its partners Fra-1 and Fra-2 in rat VSMC. Furthermore, the –138T variant form of the promoter was induced following Phorbol 12-Myristate 13-Acetate (PMA / TPA) treatment while the –138C variant was refractive to PMA treatment, confirming that AP-1 factors preferentially bind to the –138T variant. This study therefore suggests that a common polymorphism of the MGP promoter influences binding of the AP-1 complex which may lead to altered transcription and serum levels. This could have important implications for diseases such as atherosclerosis and aortic valve stenosis since it strongly suggests a genetic basis for regulation of tissue calcification.