Characterising the role of the SUMO E3-ligase PIAS in heterochromatin establishment in the Drosophila embryo
Fabe, Mathilde Thérèse Lidia
In Drosophila, position effect variegation (PEV) assays helped to uncover proteins involved in the establishment and maintenance of heterochromatin. One gene initially identified as a suppressor of variegation is Su(var)2-10 and encodes the fly PIAS ortholog (Protein inhibitor of activated STAT), a SUMO E3-ligase conserved across species. Interestingly, while PIAS and SUMO are not found at constitutive heterochromatin in somatic cells, we found it to be localised at pericentric heterochromatin during early embryogenesis. In this developmental context, we also observe that PIAS is targeted to the centromere preceding the recruitment of heterochromatin protein 1 (HP1). Further study of PIAS and SUMO interactome by co-immunoprecipitation and mass spectrometry highlighted the fly homologs of human ATRX (alpha-thalassemia and mental retardation X-linked syndrome), called XNP/dATRX and ADD1. ATRX is a chromatin remodeler of the SWI/SNF2 family and promotes incorporation of the histone variant H3.3 into chromatin. Using siRNA in early fly embryos, I could demonstrate that depletion of PIAS affected the recruitment of HP1, ADD1 and XNP to heterochromatin. Interestingly, I also observed a reduction of histone H3.3 enrichment at pericentric chromatin, suggesting that PIAS-mediated SUMOylation regulates H3.3 deposition and heterochromatin establishment in early Drosophila embryos.