Structural basis of tankyrase activation by polymerization.

The poly-ADP-ribosyltransferase tankyrase (TNKS, TNKS2) controls a wide range of disease-relevant cellular processes, including WNT-ß-catenin signalling, telomere length maintenance, Hippo signalling, DNA damage repair and glucose homeostasis1,2. This has incentivized the development of tankyrase inhibitors. Notwithstanding, our knowledge of the mechanisms that control tankyrase activity has remained limited. Both catalytic and non-catalytic functions of tankyrase depend on its filamentous polymerization3-5. Here we report the cryo-electron microscopy reconstruction of a filament formed by a minimal active unit of tankyrase, comprising the polymerizing sterile alpha motif (SAM) domain and its adjacent catalytic domain. The SAM domain forms a novel antiparallel double helix, positioning the protruding catalytic domains for recurring head-to-head and tail-to-tail interactions. The head interactions are highly conserved among tankyrases and induce an allosteric switch in the active site within the catalytic domain to promote catalysis. Although the tail interactions have a limited effect on catalysis, they are essential to tankyrase function in WNT-ß-catenin signalling. This work reveals a novel SAM domain polymerization mode, illustrates how supramolecular assembly controls catalytic and non-catalytic functions, provides important structural insights into the regulation of a non-DNA-dependent poly-ADP-ribosyltransferase and will guide future efforts to modulate tankyrase and decipher its contribution to disease mechanisms.

3.9 Å structure of the nucleosome core particle determined by phase-plate cryo-EM.

The Volta phase plate is a recently developed electron cryo-microscopy (cryo-EM) device that enables contrast enhancement of biological samples. Here we have evaluated the potential of combining phase-plate imaging and single particle analysis to determine the structure of a small protein-DNA complex. To test the method, we made use of a 200 kDa Nucleosome Core Particle (NCP) reconstituted with 601 DNA for which a high-resolution X-ray crystal structure is known. We find that the phase plate provides a significant contrast enhancement that permits individual NCPs and DNA to be clearly identified in amorphous ice. The refined structure from 26,060 particles has an overall resolution of 3.9 Å and the density map exhibits structural features consistent with the estimated resolution, including clear density for amino acid side chains and DNA features such as the phosphate backbone. Our results demonstrate that phase-plate cryo-EM promises to become an important method to determine novel near-atomic resolution structures of small and challenging samples, such as nucleosomes in complex with nucleosome-binding factors.