Molecular characterization of a heat shock cognate 70-4 promoter from the silkworm, Bombyx mori

A cDNA stretch encoding one of the silkworm HSC70 genes with its upstream sequence has been identified, based on partial cDNA sequences registered in a Bombyx mori EST database. The deduced amino acid sequence with 649 residues was 89 and 96% identical to those from Drosophila melanogaster hsc4 and Manduca sexta Hsc70-4, respectively. The expression analysis by the reverse transcription polymerase chain reaction demonstrated that the mRNA transcription occurred in all tissues examined and was not stimulated by heat shock. These results suggested that the current structure corresponds to the silkworm heat shock cognate 70-4 gene and named BmHSC70-4. Cloning and sequencing of the genomic 5'-flanking, putative promoter region of BmHSC70-4 revealed the presence of several canonical transcription elements such as the GATA box, CCAAT motif and HSE (heat shock element), although without a TATA box element. The deletion analysis of the 5' flanking region of the BmHSC70-4 gene, that fused to the firefly luciferase coding sequence as a reporter, indicated that the longest construct (-1,639/+26) showed the highest promoter activity, and the constructs -111 to -59 and -1,110 to -941 contained the basal transcription enhancer and the repressor elements, respectively.