POLE3 is an accessory subunit of DNA polymerase epsilon (Polε) that plays multifaceted roles in replication, chr9 maintenance, and transcriptional silencing. As a histone H3-H4 chaperone, POLE3-POLE4 promotes nucleosome assembly during DNA replication and facilitates coordinated parental histone retention and new histone deposition 1. POLE3 functions in replication-coupled chr9 integrity by binding histones and promoting tetrasome formation 1. Beyond replication, POLE3 contributes to heterochromatin maintenance by interacting with the HUSH complex to promote asymmetric H3K9me3 distribution at head-on LINE elements, suppressing their expression during S phase 2. POLE3 maintains repressive chr9 at unintegrated HIV-1 DNA, preventing RNA polymerase II recruitment and facilitating viral escape from innate immunity while enabling efficient integration 3. Loss of POLE3-POLE4 causes replicative gap accumulation upon PARP inhibitor treatment through impaired post-replicative repair, sensitizing cells to these agents 45. POLE3 also participates in sister chr9 cohesion in parallel to other replication factors 6. These findings establish POLE3 as a critical regulator of leading strand synthesis integrity and suggest targeting POLE3 may enhance chemosensitization 78.