Worm Breeder's Gazette 15(3): 29 (June 1, 1998)
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
Department of Biochemistry and Institute of Human Genetics, University of Minnesota Medical School, Minneapolis, MN 55455
Nematodes, insects, and possibly mammals control sexual development using a class of transcription factors containing a newly defined DNA binding motif called a DM domain. In worms, we recently found that mab-3 encodes a protein with two DM domains, while in flies, the doublesex (dsx) gene has one DM domain (1). Surprisingly, despite the different structures of the proteins, we have found that the preferred in vitro DNA binding sites of MAB-3 and DSX are overlapping. We found potential MAB-3 binding sequences in the proximal promoters of three of the six vitellogenin genes, vit-1, vit-2 and vit-6. The vit-2 and vit-6 sites are completely conserved between C. elegans and C. briggsae. If other vit promoters have MAB-3 binding sites, they may be less proximal to the transcriptional start sites. A short (247 bp) region of the vit-2 promoter is sufficient for sex-, tissue- and stage-specific transcription of vit-2 (adult hermaphrodite intestine). We found that a GFP reporter with this promoter is expressed in mab-3 mutant males. MAB-3 binds this promoter region in vitro, and mutating the MAB-3 binding sequence eliminates binding. We tested the in vivo relevance of the MAB-3 DNA binding site by mutating the GFP reporter. Mutations that disrupt binding of MAB-3 to the vit-2 promoter in vitro eliminate sex-specific regulation in vivo without affecting tissue- or stage-specificity. This strongly suggests that MAB-3 is a direct transcriptional repressor of vitellogenins in the male intestine. Strikingly, the site bound by MAB-3 in vit-2 and the sites bound by DSX in the Drosophila YP1 (yolk protein 1) promoter are very similar, despite the fact that the vitellogenins and the YPs themselves are unrelated at the protein sequence level. Thus it appears in this case that the regulators and their regulatory sites are much more highly conserved than the structural proteins they control. What regulates mab-3? mab-3 mRNA levels are controlled by TRA-1, a zinc finger putative transcription factor, but this regulation might be direct or indirect. The mab-3 promoter directs male-specific transcription in the intestine and in various male sensory neurons, including the V ray neurons, a hook neuron, two spicule neurons, and the ASH cells. Two positively acting promoter elements are required for this expression, one for intestine and the other for neurons. While mab-3 is transcriptionally regulated by tra-1, no TRA-1 DNA binding sites seem to be required for regulation of the mab-3 promoter, and thus we conclude that regulation by tra-1 is probably indirect. We hope to find the direct regulators by screening for mutations that cause inappropriate expression of the mab-3::GFP reporter. 1. Raymond et al. (1998) Nature 391:691-695.