Abstract
In Drosophila melanogaster the male specific lethal (MSL) complex is required for upregulation of expression of most X-linked genes in males, thereby achieving X chromosome dosage compensation. The MSL complex is highly enriched across most active X-linked genes with a bias towards the 3′ end. Previous studies have shown that gene transcription facilitates MSL complex binding but the type of promoter did not appear to be important. We have made the surprising observation that genes driven by the glass multiple reporter (GMR) enhancer-promoter are not dosage compensated at X-linked sites. The GMR promoter is active in all cells in, and posterior to, the morphogenetic furrow of the developing eye disc. Using phiC31 integrase-mediated targeted integration, we measured expression of lacZ reporter genes driven by either the GMR or armadillo (arm) promoters at each of three X-linked sites. At all sites, the arm-lacZ reporter gene was dosage compensated but GMR-lacZ was not. We have investigated why GMR-driven genes are not dosage compensated. Earlier or constitutive expression of GMR-lacZ did not affect the level of compensation. Neither did proximity to a strong MSL binding site. However, replacement of the hsp70 minimal promoter with a minimal promoter from the X-linked 6-Phosphogluconate dehydrogenase gene did restore partial dosage compensation. Similarly, insertion of binding sites for the GAGA and DREF factors upstream of the GMR promoter led to significantly higher lacZ expression in males than females. GAGA and DREF have been implicated to play a role in dosage compensation. We conclude that the gene promoter can affect MSL complex-mediated upregulation and dosage compensation. Further, it appears that the nature of the basal promoter and the presence of binding sites for specific factors influence the ability of a gene promoter to respond to the MSL complex.
Highlights
The Drosophila eye is well suited to genetic investigations, as it is a dispensable tissue with mutant phenotypes that are relatively easy to identify [1]
glass multiple reporter (GMR)-hid is a poor reporter of dosage compensation Our understanding of the mechanism of X chromosome dosage compensation may need to better appreciate the balance between the male specific lethal (MSL) complex and the wider network of chromatin regulators
The initial aim of this study was to search for novel dosage compensation factors, not by screening for male-lethality, but by using the reduced eye size of GMR-hid to screen for mutations that affected the end product of dosage compensation: the equalization of phenotype
Summary
The Drosophila eye is well suited to genetic investigations, as it is a dispensable tissue with mutant phenotypes that are relatively easy to identify [1]. Expression of the pro-apoptotic hid gene from GMR kills eye cells proportional to the GMR-hid expression level; a smaller eye with higher levels [7]. This reporter system facilitated a mutational screen for apoptosis-related factors, by identifying modifiers of GMR-hid-derived eye size [8]. We sought to use the variable eye size of GMR-hid to screen for modifiers of Drosophila dosage compensation, but observed that GMR-hid did not report on dosage compensation. We have investigated why GMR-hid failed to respond, and report that the minimal hsp promoter appears refractory to the compensation machinery
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