Puran S. Bora

The ethanol metabolite, linolenic acid ethyl ester, stimulates mitogen-activated protein kinase and cyclin signaling in hepatic stellate cells

Chronic ethanol consumption can result in hepatic fibrosis and cirrhosis. In addition to oxidative metabolism, ethanol can be metabolized by esterification with fatty acids to form fatty acid ethyl esters (FAEE) such as linolenic acid ethyl ester (LAEE). We have previously demonstrated that LAEE has promitogeinc and activating effects on hepatic stellate cells (HSC), but the mechanisms of these actions are not known. Intracellular signaling through MAP kinase pathways, including extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) can influence the activity of the transcription factor AP-1, while cell-cycle regulatory proteins such as cyclin E and cyclin-dependent kinase (CDK), play an important role in cell proliferation. In this study, we demonstrate that treatment of HSC with LAEE increases cyclin E expression and cyclin E/CDK2 activity, which may underlie the promitogenic effects of this compound. In addition, LAEE increases ERK and JNK activity, and these pathways play an important role in the activation of AP-1-dependent gene expression by LAEE. The stimulation of intracellular signaling pathways in HSC by this well-characterized ethanol metabolite may contribute to ethanol-induced hepatic fibrogenesis.

Human fatty acid ethyl ester synttiase-III gene: Genomic organization, nucleotide sequencing and chromosomal localization

The complete gene for human fatty acid ethyl ester Synthase-III(FAEES-III) was isolated from a human genomic lambda phage library forfunctional and structural determination. The gene spans approximately 3.3 kbwhich includes 791 base pairs of the 5′ and 124 base pairs of the 3′flanking regions. The gene is comprised of seven exons and is interrupted bysix introns. Several transcription regulatory sequences were identified inthe promoter region. Primer extension experiments demonstrated the existenceof two possible transcription initiation sites at nucleotide -29 and 32position, 5 to the start of the translation. In addition to a TATA box atposition -29 relative to the transcription initiation site and two SplGGGCGG recognition sequences at nucleotide positions -42 to -37 and -50 to-45, the promoter contains a sequence motif matching the transcriptionactivating factor AP-1. We also found an A + T rich region betweennucleotide -505 and -390 which contained twenty-two AAAAT tandem repeats.The gene for FAEES-III was localized to human chromosome 11 by hybridizingthe genomic fragment Xh01 to Chinese hamster/human somatic cell hybridpanels. These data extend our knowledge of non-oxidative alcohol metabolismand permit linkage analyses between this pathway and alcohol-relatedphenotypes.

Mutagenesis and characterization of specific residues in fatty acid ethyl ester synthase: A gene for alcohol-induced cardiomyopathy

Fatty acid ethyl ester synthase-III metabolizes both ethanol and carcinogens. Structure-function studies of the enzyme have not been performed in relation to site specific mutagenesis. In this study, three residues (Gly 32, Cys 39 and His 72) have been mutated to observe their role in enzyme activity. Gly to Gln, Cys to Trp and His to Ser mutations did not affect fatty acid ethyl ester synthase activity, but His to Ser mutant had less than 9% of control glutathione S-transferase activity. The apparent loss of transferase activity reflected a 28 fold weaker binding constant for glutathione. Thus, this study indicates that Gly and Cys may not be important for synthase or transferase activities however, histidine may play a role in glutathione binding, but it is not an essential catalytic residue of glutathione S-transferase or for fatty acid ethyl ester synthase activity.