Essential roles of Windei and nuclear monoubiquitination of Eggless/SETDB1 in transposon silencing

Eggless/SETDB1 (Egg), the only essential histone methyltransferase (HMT) in Drosophila , plays a role in gene repression, including piRNA‐mediated transposon silencing in the ovaries. Previous studies suggested that Egg is post‐translationally modified and showed that Windei (Wde) regulates Egg nuclear localization through protein–protein interaction. Monoubiquitination of mammalian SETDB1 is necessary for the HMT activity. Here, using cultured ovarian somatic cells, we show that Egg is monoubiquitinated and phosphorylated but that only monoubiquitination is required for piRNA‐mediated transposon repression. Egg monoubiquitination occurs in the nucleus. Egg has its own nuclear localization signal, and the nuclear import of Egg is Wde‐independent. Wde recruits Egg to the chromatin at target gene silencing loci, but their interaction is monoubiquitin‐independent. The abundance of nuclear Egg is governed by that of nuclear Wde. These results illuminate essential roles of nuclear monoubiquitination of Egg and the role of Wde in piRNA‐mediated transposon repression. Synopsis Eggless/SETDB1 (Egg) is a H3K9 methyltransferase essential for piRNA‐mediated transposon silencing. Its nuclear monoubiquitination and the cofactor Wde independently regulate the catalytic activity and localization of Egg. Monoubiquitination of Egg is required for its catalytic activity and occurs in the nucleus. Nuclear import of Egg depends on its own NLS, but not on the cofactor Wde. Wde regulates the nuclear levels of Egg by recruiting it to chromatin for efficient H3K9me3 deposition. Graphical Abstract Eggless/SETDB1 (Egg) is a H3K9 methyltransferase essential for piRNA‐mediated transposon silencing. Its nuclear monoubiquitination and the cofactor Wde independently regulate the catalytic activity and localization of Egg.