Giacomo Cavalli, Lauriane Fritsch, Vincent Loubière, Axelle Donjon, Julia Morales‐Sanfrutos, Eduard Sabidó, Anne‐Marie Martinez, Bernd Schuettengruber
Polycomb group (PcG) proteins are evolutionarily conserved epigenetic regulators that maintain transcriptional states during development and are frequently misregulated in disease. Here, we generated a comprehensive collection of endogenously HA-tagged alleles for the core components of Polycomb Repressive Complex 1 (PRC1) and additional chromatin regulators in Drosophila melanogaster using CRISPR/Cas9-mediated genome engineering. We show that endogenous HA tagging preserves protein expression, chromatin localization, and genome-wide binding profiles, enabling direct comparison using a common antibody to analyze distinct PcG subunits. Proteomic analyses recovered known components of canonical and non-canonical PRC1 and PRC2 complexes and identified additional PcG-associated factors. Notably, multiple components of the nuclear pore complex (NPC) were detected as PcG interactors, and genetic analyses confirmed a functional interaction between them. This resource provides a platform for standardized analysis of Polycomb function and chromatin regulation that can easily be extended to other chromatin regulatory complexes and applied across different developmental stages