Protein binding and DNase-I-hypersensitive sites in the cis-acting regulatory region of the spore-coat SP96 gene of Dictyostelium.

ABSTRACT

The spore-coat protein gene (SP96) of Dictyostelium discoideum is transcribed only in prespore cells. To identify the cis-acting region of this gene, mutant mini-genes which contained different lengths of 5' upstream region, the partially deleted SP96 coding region and ca. 600 bp of 3' flanking sequence were transformed into D. discoideum cells. Expression of the mini-genes was analysed by Northern hybridization. Our results indicate that the 5' upstream region from -686 to -494 contains an important cis-acting element for the temporal and cell type-specific transcription. A nuclear factor which specifically bound the cis-acting region was identified by gel retardation assay. DNase-I-hypersensitivity of the 5' upstream region was examined and it was shown that the appearance of two new hypersensitive sites correlates with transcriptional activation of the gene. One of the two sites maps to the TATA region and the other was located in the cis-acting region identified by deletion analysis. Our results suggest that gene activation occurs by conformational changes in the chromatin structure of the cis-acting region followed by subsequent binding of regulatory factors and the TATA-binding protein.