CPT1a over-expression increases long-chain fatty acid oxidation and reduces cell viability with incremental palmitic acid concentration in 293T cells q Ulrike L. Jambor de Sousa a , Michael D. Koss a , Marion Fillies b , Anja Gahl b , Martin R.L. Scheeder a , M. Cristina Cardoso b , Heinrich Leonhardt b,c , Nori Geary a , Wolfgang Langhans a , Monika Leonhardt a, * a Institute of Animal Sciences, Swiss Federal Institute of Technology, Zurich, Switzerland b Max Delbru ¨ ck Center for Molecular Medicine, Berlin, Germany c Department of Biology II, Ludwig Maximilians University, Munich, Germany Received 29 September 2005 Available online 13 October 2005 Abstract To test the cellular response to an increased fatty acid oxidation, we generated a vector for an inducible expression of the rate-limiting enzyme carnitine palmitoyl-transferase 1a (CPT1a). Human embryonic 293T kidney cells were transiently transfected and expression of the CPT1a transgene in the tet-on vector was activated with doxycycline. Fatty acid oxidation was measured by determining the con- version of supplemented, synthetic cis-10-heptadecenoic acid (C17:1n-7) to C15:ln-7. CPT1a over-expression increased mitochondrial long-chain fatty acid oxidation about 6-fold. Addition of palmitic acid (PA) decreased viability of CPT1a over-expressing cells in a con- centration-dependent manner. Both, PA and CPT1a over-expression increased cell death. Interestingly, PA reduced total cell number only in cells over-expressing CPT1a, suggesting an effect on cell proliferation that requires PA translocation across the mitochondrial inner membrane. This inducible expression system should be well suited to study the roles of CPT1 and fatty acid oxidation in lipotox- icity and metabolism in vivo. Ó 2005 Elsevier Inc. All rights reserved. Keywords: CPT1a; Long-chain fatty acids; Mitochondria; Lipotoxicity Carnitine palmitoyltransferase 1 (CPT1) is one of the best regulated enzymes [1]. It catalyzes the transfer of long-chain fatty acids (LCFA) from CoA to carnitine for translocation across the mitochondrial inner membrane and is well recognized as the primary rate-controlling step in fatty acid oxidation [2,3]. Many studies [4,5] indeed indi- cate that the activity of CPT1 determines the rate of LCFA oxidation and there is one study reporting that over-expres- sion of CPT1a in cultured b-cells increases fatty acid oxida- tion [6]. By regulating LCFA oxidation, CPT1 is also involved in other vital functions such as control of food in- take [7–9], insulin secretion [6], and gluconeogenesis [10,11]. In most of these studies, CPT1 activity was manip- ulated pharmacologically [9,10] with substances whose ac- tions were not wholly selective. An alternative approach is to modulate CPT1 expression by developing a transgenic animal model that permits controlled expression of CPT1 in specific organs. Given the complex interactions and redundancies in physiological control systems, however, an acutely inducible over-expression of CPT1 in specific 0006-291X/$ - see front matter Ó 2005 Elsevier Inc. All rights reserved. doi:10.1016/j.bbrc.2005.10.016 q Abbreviations: CPT, carnitine palmitoyltransferase; Dox, doxycycline; GFP, green fluorescent protein; LCFA, long-chain fatty acids; PA, palmitic acid; rtTA, Dox-inducible transactivator. * Contact author. Fax: +41 44 6557201. E-mail address: monika.leonhardt@inw.agrl.ethz.ch (M. Leonhardt). www.elsevier.com/locate/ybbrc Biochemical and Biophysical Research Communications 338 (2005) 757–761 BBRC