. The human ACAT-1 cDNA was cloned by a somatic cell and molecular genetic approach. Chinese hamster ovary (CHO) cell mutants lacking ACAT activity (including clone AC29) were isolated (8); subsequent stable transfection experiments showed that human genomic DNAs complemented the ACAT deficiency in AC29 cells (9). A 1.2-kb human genomic DNA fragment was cloned from the stable transfectants. This fragment (designated as G2 DNA) led to the eventual cloning of a full-length human ACAT cDNA K1 (4011 bp in length). Expression of this cDNA, designated as ACAT-1, in AC29 cells complemented the ACAT deficiency of the mutant (10). Additional results showed that expressing this cDNA in insect cells, which do not contain endogenous ACAT-like activity, produced high levels of ACAT activity in vitro, confirming that this cDNA encodes the catalytic component of ACAT enzyme (11). The coding region of the ACAT gene has been mapped to chromosome 1q 25 (12). Protein sequence analysis revealed the ACAT-1 protein as a hydrophobic protein containing multiple transmembrane domains and sharing several peptide regions in common with other acyltransferases (10). Recombinant human ACAT-1 protein expressed in CHO cells has been purified to homogeneity; the homogeneous ACAT-1 protein remains catalytically active and uses cholesterol as a substrate in a highly cooperative manner (13). Homologues of human ACAT-1 cDNA have also been cloned from other species (reviewed in Ref. 1), including two yeast homologues (14,15). Disruption of the ACAT-1 gene in mice has been reported (16); the ACAT-1 gene-deficient mice exhibit marked reduction in cholesteryl ester levels in only selective tissues and not in all the tissues examined. These and other results led to the molecular cloning of ACAT-2 cDNA (17-19). The predicted amino acid sequence of ACAT-2 is homologous but distinct from that of ACAT-1. The physiological roles of ACAT-1 and ACAT-2 in various tissues of different species are currently under intense investigation by several laboratories. In humans, immunodepletion experiments suggest that the ACAT-1 protein plays major catalytic roles in hepatocytes, adrenal glands, macrophages, and kidneys, but not in the intestines (20).The 4.0-kb human ACAT-1 cDNA contains a single open reading frame of 1.65 kb. It also contains an unusually long 5Ј-untranslated region (5Ј-UTR; 1396 bp) and 965 bp of 3Ј-untranslated region. Using the coding region as probe, North-

Enhanced long chain fatty acid synthesis may occur in breast cancer, where it is necessary for tumor growth and predicts a poor prognosis. ''Spot 14'' (S14) is a carbohydrate-and thyroid hormone-inducible nuclear protein specific to liver, adipose, and lactating mammary tissues that functions to activate genes encoding the enzymes of fatty acid synthesis. Amplification of chromosome region 11q13, where the S14 gene (THRSP) resides, also predicts a poor prognosis in breast tumors. We localized the S14 gene between markers D11S906 and D11S937, at the telomeric end of the amplified region at 11q13, and found that it was amplified and expressed in breast cancer-derived cell lines. Moreover, concordant expression of S14 and a key lipogenic enzyme (acetyl-CoA carboxylase) in a panel of primary breast cancer specimens strongly supported a role for S14 as a determinant of tumor lipid metabolism. S14 expression provides a pathophysiological link between two prognostic indicators in breast cancer: enhanced lipogenesis and 11q13 amplification.

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