Leukotriene C4 (
LTC4) synthase catalyzes the conjugation of
LTA4 with reduced GSH to form
LTC4, the parent of the receptor active cysteinyl
leukotrienes implicated in the pathobiology of
bronchial asthma. Previous cloning of the
cDNA for human
LTC4 synthase demonstrated significant homology of its amino acid sequence to that of
5-lipoxygenase activating
protein (FLAP) but none to that of the GSH S-
transferase super-family. Genomic cloning from a P1 library now reveals that the gene for
LTC4 synthase contains five exons (ranging from 71 to 257
nucleotides in length) and four introns, which in total span 2.52 kilobase pairs in length. The intron/exon junctions of
LTC4 synthase align identically with those of FLAP; however, the small size of the
LTC4 synthase gene contrasts with the > 31-kilobase pair size reported for FLAP. Confirmation of the
LTC4 synthase gene size to ensure that no deletions had occurred during the cloning was obtained by two overlapping polymerase chain reactions from genomic
DNA, which provided products of the predicted sizes. Primer extension analysis with
poly(A)+ RNA from culture-derived human eosinophilic granulocytes or the KG-1 myelogenous cell line revealed multiple transcriptional start sites with prominent signals at 66, 69, and 96 base pairs 5' of the ATG translation start site. The 5'-flanking region revealed a GC-rich promoter sequence consistent with an SP-1 site and consensus sequences for
AP-1 and AP-2 enhancer elements, 24, 807, and 877 bp, respectively, 5' from the first transcription initiation site. Southern blot analysis of a genomic
DNA (with full-length
cDNA as well as 5' and 3'
oligonucleotide probes) confirmed the size of the gene and indicated a single copy gene in normal human genomic
DNA. Fluorescent in situ hybridization mapped
LTC4 synthase to chromosomal location 5q35, which is in close proximity to the cluster of genes for
cytokines and receptors involved in the regulation of cells central to allergic
inflammation and implicated in
bronchial asthma.