PROMOTER STRUCTURE AND INTRON-EXON ORGANIZATION OF A SCORPION ALPHA-TOXIN GENE

The Androctonus australis scorpion venom contains alpha-toxins for whi ch the complementary DNAs have been cloned [Bougis et al, (1989) J. Bi ol. Chem. 264, 19259-19265], targeting with high affinity the voltage- sensitive sodium channel. From a genomic library made of this species of scorpion, we have cloned and characterized the gene encoding the to xin AaH I'. The gene transcriptional unit is 793 base pairs long, and the gene has a single intron of 425 base pairs located near the end of the signal peptide of the toxin precursor. The transcription initiati on site was determined by primer extension and corresponded to the nuc leotide sequence AACAA, Upstream, a promoter region has been identifie d with positive acting sequence elements at consensus positions, such as a CCAAT box and a TATA box. In addition, putative elements for bind ing the transcriptional factors MAT-alpha 2, Pit-1, and IEF1 are also present. Analysis of DNA curvature by computer modeling revealed a str ong bending centered around the transcription initiation site of the g ene. The bending angle (61 degrees) estimated experimentally using pol yacrylamide gel electrophoresis correlates well with the value predict ed by computer modeling (66 degrees). Other minor deflections of the h elix axis cooperate for an overall curvature of nearly 90 degrees, whi ch is significantly stronger than similar structures already reported in eukaryotic cells. It is worth noting that the grooves relative to t he CCAAT box and the TATA box lie along the inside of the DNA curve. T his observation is in agreement with the previously reported correlati on between DNA bending and promoter function.