Genome compaction is one of the important subject areas for understanding the mechanisms regulating genes’ expression and DNA replication and repair. The basic unit of DNA compaction in the eukaryotic cell is the nucleosome. The main chromatin proteins responsible for DNA compaction have already been identified, but the regulation of chromatin architecture is still extensively studied. Several authors have shown an interaction of ARTD proteins with nucleosomes and proposed that there are changes in the nucleosomes’ structure as a result. In the ARTD family, only PARP1, PARP2, and PARP3 participate in the DNA damage response. Damaged DNA stimulates activation of these PARPs, which use NAD⁺ as a substrate. DNA repair and chromatin compaction need precise regulation with close coordination between them. In this work, we studied the interactions of these three PARPs with nucleosomes by atomic force microscopy, which is a powerful method allowing for direct measurements of geometric characteristics of single molecules. Using this method, we evaluated perturbations in the structure of single nucleosomes after the binding of a PARP. We demonstrated here that PARP3 significantly alters the geometry of nucleosomes, possibly indicating a new function of PARP3 in chromatin compaction regulation.

DNA polymerase β (Polβ) is a central player of base excision repair (BER), performing gap-filling synthesis on damaged DNA. While nucleosome core particles (NCPs) are known to impede activity of BER enzymes, the regulation of this process in linker DNA adjacent to nucleosomes remains unclear. Here we demonstrate an unexpected stimulation of Polβ-catalyzed gap-filling and strand-displacement synthesis in linker DNA by the adjacent NCP. Notably, the nucleosomal context reinforces the regulatory modulation of Polβ activity by PARP1/PARP2 and FEN1. While linker histone H1 restricts strand-displacement synthesis at the nucleosome entry/exit site, PARP1 and PARP2 modulate Polβ function through competitive binding to DNA gaps or nicks and via poly(ADP-ribosyl)ation (PARylation). At the same time, PARPs binding differentially regulates BER sub-pathway choice, and PARylation alleviates H1-mediated inhibition. These findings reveal a multi-layered regulatory system wherein the nucleosome acts as a dynamic platform coordinating Polβ activity and its interplay with chromatin-associated factors, influencing the balance between short- and long-patch BER. The research advances understanding of chromatin-mediated control of BER DNA repair synthesis and the functional specialization of PARP1 and PARP2 in maintaining genome stability. Highlights: The nucleosome core particle acts not only as a barrier but also as a stimulator of Polβ-mediated DNA repair synthesis in adjacent linker DNA. The nucleosome acts as an allosteric platform that enhances the regulatory functions of chromatin-associated factors (PARP1, PARP2, H1) in the linker DNA repair synthesis. PARP1 suppresses overall Polβ synthesis, while PARP2 specifically inhibits strand displacement, thereby gating the choice between short- and long-patch BER pathways.

The repair processes regulation including base excision repair (BER) is implemented by a cellular signal PARylation catalysed by PARP1 and PARP2. Despite intensive studies, it is far from clear how BER is regulated by PARPs and how the roles are distributed between the PARPs. Here, we investigated the effects of PARP1, PARP2 and PARylation on activities of the main BER enzymes (APE1, Polβ and LigIIIα) in combination with XRCC1 in the nucleosomal context. We constructed nucleosomes with midward- or outward-oriented damage. It was concluded that in most cases, the presence of PARP1 leads to the suppression of the activities of APE1, Polβ, and to a lesser extent LigIIIα. PARylation by PARP1 attenuated this effect to various degrees. PARP2 had an influence predominantly on the last stage of BER: DNA sealing. Nonetheless, PARylation by PARP2 led to Polβ inhibition and to significant stimulation of LigIIIα activities in a NAD ⁺ -dependent manner.

Being the basic building blocks of chromatin, nucleosomes and their stability determine the genome accessibility for different DNA-dependent proteins. This characteristic is labile under cell-life processes. One of the abundant DNA-binding proteins, which is important for genome compaction, is poly(ADP-ribose) polymerase1 (PARP1). Despite the extensive experimental data on the chromatin compaction regulation under ADP-ribosylation, the details of the interplay of nucleosome with PARP1 in the absence of protein activation remain unclear. In this study, we discovered unusual changes of the nucleosome wrapping strength upon PARP1 interaction using a single-molecule approach—optical tweezers. We demonstrated that PARP1 binding leads to weakening of the contacts of inner DNA turn in nucleosome.