Confocal laser scanning microscopy of human skin fibroblasts showing transient expression of a green fluorescent carnitine palmitoyltransferase 1 fusion protein.

Abstract

The mitochondrial outer membrane enzyme carnitine palmitoyltransferase 1 (CPT1) is a main site of regulation of intracellular long-chain fatty acid transport. At least two isoforms of CPT1 are expressed in the body: L-CPT1 (the “liver-type” isoform) and M-CPT1 (the “muscle-type” isoform). Skin fibroblasts from healthy humans are known to contain only one isoform of CPT1: the liver-type, which is encoded by the gene CPT1A. Skin fibroblasts from patients with a liver-type CPT1 deficiency do not express either of the two known CPT1 isoforms (neither livernor muscle-type), and therefore could provide an excellent background to study CPT1 by means of molecular complementation. In this chapter, we describe the first experiments we carried out with a gene fusion of a complementary DNA of the human gene for muscle-type carnitine palmitoyltransferase (CPT1B) and a gene encoding an “enhanced” green fluorescent protein (GFP). We wished to express the human CPT1B gene in human skin fibroblasts, taking the following facts into consideration: