SET Interacts With CENP‐A and HJURP to Ensure Proper Mitotic Progression in Human Cells

Centromeres are specialized chromosomal loci that ensure faithful chromosome segregation during mitosis, a function that depends on the precise localization of the histone H3 variant centromere protein A (CENP-A) within centromeric chromatin. CENP-A deposition is tightly regulated by assembly factors, including HJURP; however, the complete regulatory network remains incompletely defined. Using CENP-A immunoprecipitation coupled with mass spectrometry, we identified SET, a multifunctional chromatin-associated protein involved in transcription and nucleosome assembly, as one of the most enriched proteins in CENP-A immunoprecipitates, suggesting an association between SET and CENP-A. Here, we investigated this association and its functional significance. Reciprocal co-immunoprecipitation assays showed that SET associates with both CENP-A and HJURP. Immunofluorescence analysis revealed that SET localizes to centromeres and that SET depletion reduces centromeric CENP-A enrichment. Live-cell imaging further showed that SET depletion causes chromosome alignment defects and prolonged mitotic arrest. Together, these findings identify SET as a previously unrecognized regulator of the CENP-A-HJURP axis and suggest that SET supports proper CENP-A localization at centromeres and faithful mitotic progression. This study expands our understanding of the regulatory network that maintains centromere identity.