Cloning and sequence analysis of the human acidic fibroblast growth factor gene and its preservation in leukemia patients

Abstract

Acidic fibroblast growth factor (aFGF), also known as heparin-binding growth factor 1, is a mitogen for a variety of mesoderm- and neuroectoderm-derived cells. Several different aFGF mRNA species resulting from alternative splicing have been reported. These results suggest that the gene structure and regulatory mechanism for gene expression of aFGF are complex. As a first step toward understanding aFGF gene structure, we have isolated nine overlapping genomic DNA clones spanning 54 kbp and determined the complete DNA sequences of all three coding exons. Comparison of the nucleotide sequences between the human and bovine DNA showed that the sequence similarity extended 2400 bp downstream from the coding region. Cloning of the aFGF gene allowed us to characterize this locus in acute nonlymphocytic leukemia (ANLL) patients. A fraction of ANLL patients (10-20%) have a deletion in the long arm of chromosome 5, whose distal breakpoint overlaps the aFGF locus. Therefore, a prospective cohort of eight ANLL patients was screened using three different repetitive sequence-free probes derived from the aFGF locus. Using beta-globin gene as a normalization probe for hybridizing band intensities, we conclude that there is no allelic loss or gross rearrangement within the 40 kbp stretch of the aFGF gene locus in ANLL patients with or without 5q- deletion. Consistent with this observation, the aFGF mRNA was not detected in the mononuclear cells derived from either an ANLL patient or a normal individual as judged by the reverse transcription and polymerase chain reaction. We also identified a DNA fragment, 10.7 kbp upstream from the first coding exon of human aFGF, whose sequence is conserved in both the primate and rodent genomes. Further characterization of this fragment is likely to provide insight into the significance of this high degree of conservation.