STRUCTURAL FEATURES OF THE HUMAN GENE FOR MUSCLE-SPECIFIC ENOLASE - DIFFERENTIAL SPLICING IN THE 5'-UNTRANSLATED SEQUENCE GENERATES 2 FORMSOF MESSENGER-RNA

We report here the isolation and characterization of the human gene fo r the beta or muscle-specific isoform of the glycolytic enzyme enolase . The nucleotide sequence analysis revealed structural features, such as organization as 11 coding exons, the first exon consisting of an un translated sequence and hence resembling sequences of the other two me mbers of the gene family, the alpha and gamma enolase genes. The beta enolase locus spans about 6 kbp genomic DNA. Sequences matching the co nsensus sequence for muscle-specific regulatory factors are present in the 5'-flanking region and within the first intron. A combination of primer extension, S1 nuclease protection and RNA-sequencing experiment s indicates that the gene has a unique transcriptional start site, 26 bp downstream of a TATA-like box; the differential usage of two donor sites within the untranslated exon I generates two alternatively splic ed transcripts. The existence of the two mRNA, differing from one anot her in the presence or absence of a 42-nucleotide fragment in the lead er sequence, was confirmed by cloning the corresponding cDNA using the rapid amplification of cDNA ends strategy. Secondary-structure predic tions indicated that the leader sequences of the spliced forms could f orm hairpin structures with different free energies of formation, sugg esting translational control.