Faculty Bibliography

Signalling by steroid hormones is mediated by receptor proteins that bind hormonal ligands and regulate the transcription of specific genes. The heat-shock protein hsp90 seems to associate selectively with unliganded receptors (aporeceptors), but it has not been determined whether this interaction affects receptor function in vivo. To address the role of hsp90, we have taken advantage of the capacity of mammalian steroid receptors to function in yeast. We constructed a strain of Saccharomyces cerevisiae in which hsp90 expression was regulatable and could be reduced more than 20-fold relative to wild type. At low levels of hsp90, aporeceptors seem to be mostly hsp90-free, yet fail to enhance transcription; on hormone addition, the receptors are activated but with markedly reduced efficiency. Thus hsp90 does not inhibit receptor function solely by steric interference; rather, hsp90 seems to facilitate the subsequent response of the aporeceptor to the hormonal signal. This is the first biological evidence that hsp90 acts in the signal transduction pathway for steroid receptors

Yolk protein genes 1 and 2 (yp1 and yp2) of Drosophila melanogaster are divergently transcribed neighboring genes. Both are transcribed in only two tissues, the ovarian follicle cells and the fat bodies of adult females. Previous work has identified a yolk protein enhancer between the genes that is sufficient to direct transcription in one of the tissues, female fat bodies. Using germ-line transformation methods, we identify two cis-acting regions with positive effects on transcription in ovaries. One, a 301-bp region located between the genes, influences both genes and is an enhancer determining the stage and cell type specificity of ovarian transcription. The other, a 105-bp region located in the first exon of yp2, acts across the yp2 promoter region to stimulate yp1 transcription in ovaries. Additional observations suggest how a single enhancer influences both promoters

The entire sequence of the Drosophila melanogaster yolk protein 3 (YP3) gene (yp3), including 1822 nucleotides (nt) of 5'- and 834 nt of 3'-flanking DNA, has been determined. In addition, the 5' and 3' ends of the mRNA and the two introns have been mapped. The predicted amino acid sequence of YP3 has considerable homology (43%) to the other two yolk proteins of D. melanogaster. The nucleotide sequence of yp3 was compared to the other two yolk protein genes which have the same developmental pattern of expression. In addition to extensive homology between the protein coding regions, we found two small regions of homology between yp3 flanking sequences and a segment of DNA required for normal expression of the yolk protein 1 gene in adult female fat bodies

The yolk protein 1 gene (yp1) of Drosophila melanogaster is expressed only in the ovarian follicle cells and the fat bodies of adult females. We have previously shown that a different cis-acting DNA region is required for each of these tissue specificities. In this paper we use germ line transformation to localize and characterized one of these tissue-specific regulatory regions. We demonstrate that a 125 bp segment of DNA located 196 bp upstream of the yp1 cap site is sufficient to determine the sex-, stage-, and fat body-specific expression of the yp1 gene. We also find that this region can confer yp1-specific expression on a heterologous Drosophila promoter. This specificity is retained when the region is in different orientations and at different distances from the heterologous promoter. Thus a small regulatory region acts in vivo as a positive enhancer to determine the fat body expression pattern of yp1

The adjacent and divergently transcribed yp1 and yp2 genes of Drosophila were separated at a site that is 342 base pairs upstream of yp2 and 883 base pairs upstream of yp1. Each gene was separately used to transform Drosophila germ-line-producing flies with a single copy of the introduced gene. Transcripts from the introduced genes were found only in adult females. Thus, the introduced genes maintain the sex- and time-specific expression pattern of the endogenous yp genes. However, the pattern of tissue-specific expression differed among the two introduced genes and the endogenous genes. Transcripts from both of the endogenous genes are found in fat bodies and ovaries. In contrast, transcripts from the introduced yp1 gene are found only in fat bodies, and those from the introduced yp2 gene are found only in ovaries. Thus, each introduced DNA segment lacks at least one of the cis-acting elements required for the normal pattern of tissue-specific expression. These results indicate that the expression of a yp gene in different tissues is determined by different cis-acting elements

In this paper we have demonstrated that at least two tissue-specific cis-acting DNA elements are necessary for the normal developmental pattern of Drosophila yp1 gene expression. One of these elements is necessary for expression in the adult female follicle cells and the other is necessary for expression in adult female fat bodies. We have localized the fat body expression element to a 125-bp fragment that lies between nucleotides -196 and -321 of the yp1 gene. This small fragment has at least one of the characteristics of enhancer sequences because it can direct a heterologous Drosophila promoter to be transcribed with the yp1 fat body expression pattern