1) Structure, Interactions and Inhibition of (poly)-ADP-ribosyl-polymerases (PARP)
PARP1 and PARP2 are abundant nuclear enzymes that act as chromatin architectural proteins, thereby shaping chromatin structure. Upon encountering DNA damage, they become enzymatically active and catalyzes the polymerization of long chains of (poly)-ADP-ribose to many target proteins, including histones, in response to DNA damage. Inhibitors of PARP1 are effective to treat many cancers.
We are interested in how PARP1 moves through DNA in vitro and in the cell (Bowerman et al., 2022; Mahadevan, Bowerman, et al., 2019; Mahadevan, Rudolph, et al., 2019; Rudolph et al., 2018), and we have arrived at a much better understanding of PARP1 enzymology and PARP inhibition ((Lin et al., 2022; Nakamoto et al., 2019; Rudolph et al., 2022; Rudolph, Roberts, & Luger, 2021; Rudolph, Roberts, Muthurajan, et al., 2021), with the goal of developing better inhibitors of PARP to treat cancer. Through cryoEM we have made progress in understanding how PARPs interact with chromatin (Gaullier et al., 2020; Rudolph, Muthurajan, et al., 2021). The auxiliary protein Histone Parylation Factor profoundly affects
PARP1 enzymatic properties and its interaction with Parp inhibitors (used for cancer therapies) (Rudolph et al., 2022; Rudolph, Roberts, & Luger, 2021; Rudolph, Roberts, Muthurajan, et al., 2021; Stojanovic et al., 2023). We use a combination of mechanistic enzymology, structural biology, and molecular biology and life-cell imaging to understand how PARP1 acts in healthy and damaged cells, and how inhibition can improve cancer therapies.
Figure legend: PARP1 interacts with nucleosomes via its BRCT domain. From Rudolph et al., 2021
2) Chaperoning Histones and Nucleosomes
Nucleosomes present profound obstacles to the machinery that requires access to the genome. The histone chaperone FACT (a key player in all DNA-dependent processes) engages and protects partially disassembled nucleosomes (Liu et al., 2020; Wang et al., 2018). We have detailed the structure and nucleosome assembly mechanism of the ATP-dependent chromatin remodeler SMARCAD1 (Markert et al., 2021).
We have also identified an unexpected role of the centromeric protein CENP-N in promoting unique chromatin structures by ‘decoding’ and compacting centromeric nucleosomes (Pentakota et al., 2017; Zhou et al., 2022). 
Figure legend: FACT recognizes and restores partially unwrapped nucleosomes From Liu et al, 2020
3) Nucleosomes for all: Histones in non-eukaryotic organisms
Histones are amongst the most conserved proteins in eukaryotes, and as such nucleosome structure and the elaborate machinery that deals with it is highly conserved. We are interested in the evolutionary origins of the nucleosome, which we probe by investigating histone-based DNA organization in non-eukaryotic organisms. We have demonstrated the existence of nucleosome-like structures in giant viruses (Liu et al., 2021) and archaea (Mattiroli et al., 2017) (Bowerman et al., 2021) (Laursen et al., 2020). Nucleosomes from giant viruses are composed of fused histone doublets with only weak sequence homology to the core histones of Amoebae (their host), and they are essential for viral fitness. In contrast, archaeal histones form slinky-like assemblies from a single histone, and these can open stochastically to permit access to the DNA in the cell.
Surprisingly, select bacteria also encode histones. We have recently discovered that histones from the predatory bacterium Bdellovibrio bacteriovorus reverse common histone paradigms by coating a straight DNA fragment rather than wrapping around it (Hocher et al., 2023). 
Figure legend: Histone-based chromatin in non-eukaryotic organisms. Right: a single archaeal histone organizes DNA into dynamic ‘slinkies’ that open and close stochastically (Mattiroli et al., 2017) (Bowerman et al., 2021). Top: Giant viruses form destabilized nucleosomes with fused histone doublets, but otherwise resemble eukaryotic nucleosomes (top right) (Liu et al., 2021). left: a bacterial histone encases DNA in a nucleohistone filament (Hocher et al., 2023).
Bowerman, S., Mahadevan, J., Benson, P., Rudolph, J., & Luger, K. (2022). Automated modeling of protein accumulation at DNA damage sites using qFADD.py [Software Report]. Biological Imaging, 2(e8). https://doi.org/https://doi.org/10.1017/S2633903X22000083
Bowerman, S., Wereszczynski, J., & Luger, K. (2021). Archaeal chromatin ‘slinkies’ are inherently dynamic complexes with deflected DNA wrapping pathways. Elife, 10. https://doi.org/10.7554/eLife.65587
Gaullier, G., Roberts, G., Muthurajan, U. M., Bowerman, S., Rudolph, J., Mahadevan, J., . . . Luger, K. (2020). Bridging of nucleosome-proximal DNA double-strand breaks by PARP2 enhances its interaction with HPF1. PLoS One, 15(11), e0240932. https://doi.org/10.1371/journal.pone.0240932
Hocher, A., Laursen, S. P., Radford, P., Tyson, J., Lambert, C., Stevens, K. M., . . . Warnecke, T. (2023). Histones with an unconventional DNA-binding mode in vitro are major chromatin constituents in the bacterium Bdellovibrio bacteriovorus. Nat Microbiol, 8(11), 2006-2019. https://doi.org/10.1038/s41564-023-01492-x
Laursen, S. P., Bowerman, S., & Luger, K. (2020). Archaea: the final frontier of chromatin. J Mol Biol, 166791. https://doi.org/10.1016/j.jmb.2020.166791
Lin, X., Jiang, W., Rudolph, J., Lee, B. J., Luger, K., & Zha, S. (2022). PARP inhibitors trap PARP2 and alter the mode of recruitment of PARP2 at DNA damage sites. Nucleic Acids Res, 50(7), 3958-3973. https://doi.org/10.1093/nar/gkac188
Liu, Y., Bisio, H., Toner, C. M., Jeudy, S., Philippe, N., Zhou, K., . . . Luger, K. (2021). Virus-encoded histone doublets are essential and form nucleosome-like structures. Cell, 184(16), 4237-4250 e4219. https://doi.org/10.1016/j.cell.2021.06.032
Liu, Y., Zhou, K., Zhang, N., Wei, H., Tan, Y. Z., Zhang, Z., . . . Luger, K. (2020). FACT caught in the act of manipulating the nucleosome. Nature, 577(7790), 426-431. https://doi.org/10.1038/s41586-019-1820-0
Mahadevan, J., Bowerman, S., & Luger, K. (2019). Quantitating repair protein accumulation at DNA lesions: Past, present, and future. DNA Repair (Amst), 102650. https://doi.org/10.1016/j.dnarep.2019.102650
Mahadevan, J., Rudolph, J., Jha, A., Tay, J. W., Dragavon, J., Grumstrup, E. M., & Luger, K. (2019). Q-FADD: A Mechanistic Approach for Modeling the Accumulation of Proteins at Sites of DNA Damage. Biophys J, 116(11), 2224-2233. https://doi.org/10.1016/j.bpj.2019.04.032
Markert, J., Zhou, K., & Luger, K. (2021). SMARCAD1 is an ATP-dependent histone octamer exchange factor with de novo nucleosome assembly activity. Sci Adv, 7(42), eabk2380. https://doi.org/10.1126/sciadv.abk2380
Mattiroli, F., Bhattacharyya, S., Dyer, P. N., White, A. E., Sandman, K., Burkhart, B. W., . . . Luger, K. (2017). Structure of histone-based chromatin in Archaea. Science, 357(6351), 609-612. https://doi.org/10.1126/science.aaj1849
Nakamoto, M. Y., Rudolph, J., Wuttke, D. S., & Luger, K. (2019). Nonspecific Binding of RNA to PARP1 and PARP2 Does Not Lead to Catalytic Activation. Biochemistry, 58(51), 5107-5111. https://doi.org/10.1021/acs.biochem.9b00986
Pentakota, S., Zhou, K., Smith, C., Maffini, S., Petrovic, A., Morgan, G. P., . . . Luger, K. (2017). Decoding the centromeric nucleosome through CENP-N. Elife, 6. https://doi.org/10.7554/eLife.33442
Rudolph, J., Jung, K., & Luger, K. (2022). Inhibitors of PARP: Number crunching and structure gazing. Proc Natl Acad Sci U S A, 119(11), e2121979119. https://doi.org/10.1073/pnas.2121979119
Rudolph, J., Mahadevan, J., Dyer, P., & Luger, K. (2018). Poly(ADP-ribose) polymerase 1 searches DNA via a ‘Monkey Bar’ mechanism. Elife, 7. https://doi.org/10.7554/eLife.37818
Rudolph, J., Muthurajan, U. M., Palacio, M., Mahadevan, J., Roberts, G., Erbse, A. H., . . . Luger, K. (2021). The BRCT domain of PARP1 binds intact DNA and mediates intrastrand transfer. Mol Cell, 81(24), 4994-5006 e4995. https://doi.org/10.1016/j.molcel.2021.11.014
Rudolph, J., Roberts, G., & Luger, K. (2021). Histone Parylation factor 1 contributes to the inhibition of PARP1 by cancer drugs. Nat Commun, 12(1), 736. https://doi.org/10.1038/s41467-021-20998-8
Rudolph, J., Roberts, G., Muthurajan, U. M., & Luger, K. (2021). HPF1 and nucleosomes mediate a dramatic switch in activity of PARP1 from polymerase to hydrolase. Elife, 10. https://doi.org/10.7554/eLife.65773
Stojanovic, P., Luger, K., & Rudolph, J. (2023). Slow Dissociation from the PARP1-HPF1 Complex Drives Inhibitor Potency. Biochemistry. https://doi.org/10.1021/acs.biochem.3c00243
Wang, T., Liu, Y., Edwards, G., Krzizike, D., Scherman, H., & Luger, K. (2018). The histone chaperone FACT modulates nucleosome structure by tethering its components. Life Sci Alliance, 1(4), e201800107. https://doi.org/10.26508/lsa.201800107
Zhou, K., Gebala, M., Woods, D., Sundararajan, K., Edwards, G., Krzizike, D., . . . Luger, K. (2022). CENP-N promotes the compaction of centromeric chromatin. Nat Struct Mol Biol, 29(4), 403-413. https://doi.org/10.1038/s41594-022-00758-y