Drosophila Rumpelstiltskin regulates P olycomb cis and trans interactions and Homie barrier activity

Background Three-dimensional genome organization is maintained in part by chromatin insulators, DNA–protein complexes that create boundaries between active and inactive chromatin and regulate enhancer-promoter interactions. In Drosophila, the Homing insulator at eve (Homie) sequence prevents repressive Polycomb (PcG) chromatin at even-skipped ( eve ) from spreading into the neighboring essential gene, TER94 . The exact mechanisms controlling Homie barrier activity are understudied. Results Here, use of an in vivo reporter assay identified the requirement of the hnRNP M homolog, Rumpelstiltskin (Rump), for Homie barrier activity. Interestingly, Rump depletion resulted in no changes to chromatin insulator protein binding at Homie but extensive chromatin association changes to the Polycomb response element (PRE)-associated protein Crooked legs (Crol) at Crol-bound PREs. Furthermore, global chromatin association changes were detected for a subset of Polycomb repressive complex 1 and 2 (PRC1 and PRC2) components. Finally, we observed increased cis- compaction at Polycomb domains genome-wide including eve and increased distances between eve and other Polycomb domains in trans after Rump depletion. Conclusions These data identify Rump as a novel regulator of Homie barrier activity and suggest a regulatory role of Rump for both cis and trans interactions of PcG proteins. Importantly, this regulatory role likely helps to maintain Homie barrier activity independently of chromatin insulator proteins.