The Clustered Regularly Interspaced Palindromic Repeats (CRISPR) system is an adaptive immune system in prokaryotes. Interference complexes encoded by CRISPR-associated (cas) genes utilize small RNAs for homology-directed detection and subsequent degradation of invading genetic elements, and they have been classified into three main types (I–III). Type III complexes share the Cas10 subunit but are subclassifed as type IIIA (CSM) and type IIIB (CMR), depending on their specificity for DNA or RNA targets, respectively. The role of CSM in limiting the spread of conjugative plasmids in Staphylococcus epidermidis was first described in 2008. Here, we report a detailed investigation of the composition and structure of the CSM complex from the archaeon Sulfolobus solfataricus, using a combination of electron microscopy, mass spectrometry, and deep sequencing. This reveals a three-dimensional model for the CSM complex that includes a helical component strikingly reminiscent of the backbone structure of the type I (Cascade) family.

Poly (ADP-ribose) polymerases (PARP) 1-3 are well-known multi-domain enzymes, catalysing the covalent modification of proteins, DNA, and themselves. They attach mono- or poly-ADP-ribose to targets using NAD⁺ as a substrate. Poly-ADP-ribosylation (PARylation) is central to the important functions of PARP enzymes in the DNA damage response and nucleosome remodelling. Activation of PARP happens through DNA binding via zinc fingers and/or the WGR domain. Modulation of their activity using PARP inhibitors occupying the NAD⁺ binding site has proven successful in cancer therapies. For decades, studies set out to elucidate their full-length molecular structure and activation mechanism. In the last five years, significant advances have progressed the structural and functional understanding of PARP1-3, such as understanding allosteric activation via inter-domain contacts, how PARP senses damaged DNA in the crowded nucleus, and the complementary role of histone PARylation factor 1 in modulating the active site of PARP. Here, we review these advances together with the versatility of PARP domains involved in DNA binding, the targets and shape of PARylation and the role of PARPs in nucleosome remodelling.

Recognition of DNA double‐strand breaks during non‐homologous end joining is carried out by the Ku70–Ku80 protein, a 150 kDa heterodimer that recruits the DNA repair kinase DNA‐dependent protein kinase catalytic subunit (DNA‐PKcs) to the lesion. The atomic structure of a truncated Ku70–Ku80 was determined; however, the subunit‐specific carboxy‐terminal domain of Ku80—essential for binding to DNA‐PKcs—was determined only in isolation, and the C‐terminal domain of Ku70 was not resolved in its DNA‐bound conformation. Both regions are conserved and mediate protein–protein interactions specific to mammals. Here, we reconstruct the three‐dimensional structure of the human full‐length Ku70–Ku80 dimer at 25 Å resolution, alone and in complex with DNA, by using single‐particle electron microscopy. We map the C‐terminal regions of both subunits, and their conformational changes after DNA and DNA‐PKcs binding to define a molecular model of the functions of these domains during DNA repair in the context of full‐length Ku70–Ku80 protein.