The balance between fatty acid (FA) catabolism and anabolism plays a central role in obesity and obesity-related complications; and understanding derangements in metabolism that underlie disease will aid in setting strategies for treating disease. The activation of long chain FA for cellular metabolism requires the five mammalian long chain acyl-CoA synthetases (ACSL) that catalyze the conversion of long chain FA into their acyl-CoA derivatives. The reason mammals need five unique ACSL isoenzymes remains unknown. The purpose of this study was to determine the role of ACSL one (ACSL1) in FA metabolism in adipose tissue and heart. To study ACSL1 in adipose, we generated an adipose-specific ACSL1 knockout mouse, the Acsl1A-/- mouse. ACSL1 in adipose was believed to be essential for the synthesis of triacylglycerol. However, in Acsl1A-/- white and brown adipocytes, the rate of TAG synthesis was similar to controls, whereas FA oxidation in isolated adipocytes and mitochondria was reduced 50-90%. Acsl1A-/- mice had increased fat mass and were severely cold intolerant. Their reduced adipose FA oxidation and marked cold intolerance indicate that normal activation of FA for oxidation in adipose tissue in vivo requires ACSL1. To study ACSL1 in heart, we generated a multi-tissue temporally induced ACSL1 knockout mouse, Acsl1T-/-. Although cardiac ACSL1 is the most abundant of the ACSL isoenzymes, its role in cardiac FA metabolism had remained unclear. In Acsl1T-/- mice, acyl-CoA synthetase activity was reduced ~90%, acyl-CoA content was reduced 65%, and long-chain acyl-carnitine species and palmitate oxidation were 80-90% lower than in control hearts. Acsl1T-/- hearts developed hypertrophy, increased mitochondrial content, and had 5-fold greater phosphorylation of S6 kinase, a target of mTOR kinase. These data suggest that ACSL1 catalyzes the initial step in the pathway of heart FA oxidation and that without ACSL1, diminished FA oxidative capacity leads to mTOR activation and results in cardiac hypertrophy and increased mitochondria. Together, these data suggest that ACSL1 has a specific function in directing the metabolic partitioning of FA towards [beta]-oxidation in both adipose tissue and heart. This dissertation has elucidated the novel role for ACSL1 as the first and required step for the oxidation of FA.