Localization of the hsp83 transcript within a 3292 nucleotide sequence from the 63B heat shock locus ofD. melanogaster

Abstract

We have determined the complete nucleotide sequence of a 3292 bp cloned segment derived from the 63B heat shock cytogenetic locus of D. melanogaster. Within this segment we have positioned the start of transcription and RNA splice sites of the unique gene that encodes the 83,000 d heat shock polypeptide (hsp83 gene) by S1 mapping and synthesis of cDNA from restriction fragment primed mRNA. The sequence begins at a point 879 bp upstream from the transcription start and includes the 149 bp nontranslated first exon, the 1139 bp intron and extends 1125 bp into the protein coding region. These data identify a single open translation reading frame for the first 375 amino acids of the 83,000 d polypeptide, beginning with the first ATG codon located at the 3′ intron-exon junction. We discuss and demonstrate the use of E. coil exonuclease III generated single-strand DNA probes as an alternative to strand separation for S1 mapping of mRNA. We also use homology search criteria based upon known protein-DNA binding sites to compare our hsp83 sequence with other sequenced Drosophila heat shock genes. These comparisons indicate that a large region of approximately 80 bp centered around the transcription initiation point of the hsp83 gene shares only a 31% homology with the corresponding region of the hsp70 gene, whereas the hsp22, 23, 26, and 27 genes 8hare a 54% homology with hsp70 in this region. The lower homology of the hsp83 gene is consistent with the deviant nature of this heat shock gene.