Supplementary Material for: Interaction Study of the Male Specific Lethal (MSL) Complex and Trans-Acting Dosage Effects in Metafemales of <i>Drosophila melanogaster</i> X.Sun J.A.Birchler 2009 The effect of ectopic expression of male specific lethal 2 <i>(msl2)</i> on chromatin modification and gene expression was studied in Drosophila diploid females and metafemales (3X;2A). Results show that ectopic expression of MSL2 in transgenic <i>msl2</i> females and metafemales sequesters the MOF histone acetylase to the X, which occurs concordantly with an increase of histone acetylation. Gene expression studies indicate that the X-linked genes are not affected by direct targeting of the MSL complex and the resulting increased H4Lys16 acetylation on the X chromosomes, suggesting one function of the MSL complex is to nullify the effect of a high level of histone acetylation. These results are not consistent with the hypothesis that the presence of the MSL complex conditions a two-fold upregulation. Autosomal gene expression is generally decreased in ectopically expressed MSL2 females, which correlates with the reduced autosomal histone acetylation. Metafemales show dosage compensation of X-linked genes with some autosomal reductions in expression. Interestingly, in metafemales with ectopically expressed MSL2, the autosomal expression is returned to a more normal level. There is a lower autosomal level of histone acetylation compared to the normal metafemales, suggesting a nullifying effect on the negative dosage effect of the X chromosome as previously hypothesized to occur in normal males.