Domains of HP1 Required for Drosophila melanogaster Heterochromatin Spreading

Centric regions of eukaryotic genomes are packaged into heterochromatin, which possesses the ability to spread along the chromosome and silence gene expression. The process of spreading has been challenging to study at the molecular level due to repetitious sequences within centric regions. An HP1 tethering system was developed that generates “ectopic heterochromatin” at sites within euchromatic regions of the Drosophila melanogaster genome. Using this system, we show that HP1 dimerization and the PxVxL interaction platform formed by dimerization of the HP1 chromo shadow domain are necessary for spreading to a downstream reporter gene located 3.7 kb away. Surprisingly, either the HP1 chromo domain or the chromo shadow domain alone is sufficient for spreading and silencing at a downstream reporter gene located 1.9 kb away. Spreading is dependent on at least two H3K9 methyltransferases, with SU(VAR)3-9 playing a greater role at 3.7 kb reporter and dSETDB1 predominately acting at 1.9 kb. These data support a model whereby HP1 takes part in multiple mechanisms of silencing and spreading. INTRODUCTION Heterochromatin Protein 1 (HP1) was identified in Drosophila as a non-histone chromosomal protein enriched in centric heterochromatin (JAMES et al. 1989; JAMES and ELGIN 1986). On polytene chromosomes, HP1 localizes near centromeres and telomeres, along the fourth chromosome and at ~200 sites within the euchromatic arms (FANTI et al. 2003; JAMES et al. 1989). Heterochromatin has the ability to “spread”, or propagate in cis, along the chromosome (WEILER and WAKIMOTO 1995). Spreading is observed when a chromosomal rearrangement