Identification of Chromatin-Associated Regulators of MSL Complex Targeting in Drosophila Dosage Compensation

Abstract

Sex chromosome dosage compensation in Drosophila provides a model for understanding how chromatin organization can modulate coordinate gene regulation. Male Drosophila increase the transcript levels of genes on the single male X approximately two-fold to equal the gene expression in females, which have two X-chromosomes. Dosage compensation is mediated by the Male-Specific Lethal (MSL) histone acetyltransferase complex. Five core components of the MSL complex were identified by genetic screens for genes that are specifically required for male viability and are dispensable for females. However, because dosage compensation must interface with the general transcriptional machinery, it is likely that identifying additional regulators that are not strictly male-specific will be key to understanding the process at a mechanistic level. Such regulators would not have been recovered from previous male-specific lethal screening strategies. Therefore, we have performed a cell culture-based, genome-wide RNAi screen to search for factors required for MSL targeting or function. Here we focus on the discovery of proteins that function to promote MSL complex recruitment to “chromatin entry sites,” which are proposed to be the initial sites of MSL targeting. We find that components of the NSL (Non-specific lethal) complex, and a previously unstudied zinc-finger protein, facilitate MSL targeting and display a striking enrichment at MSL entry sites. Identification of these factors provides new insight into how MSL complex establishes the specialized hyperactive chromatin required for dosage compensation in Drosophila.

Author Summary

Gene regulation is essential to all living things. For example, levels of gene expression in individual cells must be fine-tuned during development and in response to changing environmental conditions. Genes are regulated by DNA binding proteins and by factors that influence DNA packaging into chromatin. The MSL complex in Drosophila melanogaster is a chromatin-modifying complex that specifically regulates a large number of genes. The MSL complex targets active genes on the single male X chromosome to upregulate their output to match both female X chromosomes. How the MSL complex specifically targets the X chromosome and upregulates active genes is only partially understood. In order to increase our understanding of gene regulation at a mechanistic level, we performed a genome-wide genetic screen in male cells to identify factors that facilitate MSL targeting and function. Our results identify two chromatin-associated protein complexes and a new candidate DNA binding protein as key factors in MSL–based regulation. We also provide an extensive list of additional candidate genes to be examined in future studies.