Diamond Light Source - Annual Review 2022/23 - Concise Edition

20 21 D I A M O N D L I G H T S O U R C E A N N U A L R E V I E W 2 0 2 2 / 2 3 D I A M O N D L I G H T S O U R C E A N N U A L R E V I E W 2 0 2 2 / 2 3 Biological Cryo-Imaging Group T he Biological Cryo-Imaging Group brings together dedicated facilities for X-ray, light, and electronmicroscopy at Diamond Light Source. The electron Bio-Imaging Centre (eBIC) is the national centre for Cryo-Electron Microscopy (cryo-EM) in the UK and provides a range of capabilitiesandsupportingfacilitiesforcryo-EMandCorrelativeLightandElectronMicroscopy(CLEM).BeamlineB24hostsafullfieldcryo- transmission X-ray microscope dedicated to biological X-ray imaging and has also established a cryo super resolution fluorescence microscopy facility, which is a joint venture between Diamond and the University of Oxford. It provides a unique platform for correlative light and X-ray microscopy, and cryo-EM. A recent external beamline review rated the B24 facility as excellent and world leading. In particular, the panel commended the beamline team on establishing an internationally unique correlative platform combining two high-end 3D cryomicroscopy techniques (Cryo Soft X-ray Tomography (Cryo-SXT) and Cryo Structured IlluminationMicroscopy (Cryo-SIM) with user friendly protocols. Recent studies on B24 and eBIC this year include those to understand modifications in cells infected by the Herpes virus, insights into filaments of Parkinson’s disease and determining the structure of key proteins involved in many cellular process relevant to cancer Herpes simplex virus can rearrange the contents of human cells Herpes simplex virus (HSV)-1 is a highly prevalent human pathogen that usually manifest as cold sores or genital herpes. However, HSV-1 infection can cause life-threatening disease, and peoplewithweakened immune systems such as neonates, the elderly and patients on certain immunosuppressive drugs tend to be more at risk. HSV-1 has also been developed as a potential therapeutic; it is already in use as an anti-cancer agent (T-VEC) and is under development as a delivery vector for gene therapy. Researchers from the University of Cambridge sought to image infected cells that are as close as possible to how they would look in the cellular context. They used the facilities on Diamond’s B24 beamline to flash-freeze infected cells in liquid ethane, cryogenically preserving them in a‘near-native’state, and to perform cryo-soft-X-ray tomography (cryo-SXT). The cryo-SXT analysis allowed them to reconstruct 3D images of infected cells at very high resolution. Combining this analysis with a special fluorescent ‘timestamp’ strain of HSV- 1, constructed in Cambridge, allowed them to work out the stage of infection each cell was in when it was frozen. They were able to identify individual virus particles within the infected cells and to see how virus infection progressively changes the shape and distribution of celullar components and organelles. This study demonstrates the power of cryo-SXT for monitoring virus infection and highlights which organelles to focus on as we study the molecular characteristics of herpes virus infection. Nahas, KL. et al. DOI: 10.1371/journal. ppat.1010629 The atomic structure of alpha-synuclein filaments of Parkinson’s disease Parkinson’s disease is a neurodegenerative disorder that affects millions of people around the world. Although Parkinson’s disease is most commonly diagnosed in older adults, it can also affect younger people. In many neurodegenerative diseases, one or a few different proteins form aggregates, i.e. amyloidfilaments, in thebrain. InParkinson’s disease, theamyloid filaments aremade of the protein alpha-synuclein. However, the structures of the filaments from Parkinson’s disease remained unknown. One of the difficulties was that many of them do not twist, which leads to technical problems in the structure determination process. An international team of researchers solved the structure of alpha-synuclein filaments. They collected many electron microscope images at eBIC and sifted through them all to find the minority of the filaments that did twist. Knowledge of the structure of the alpha-synuclein filaments can be used to develop new molecules that could be useful in the clinic. For example, it may now be possible to use structure-based design to develop small molecules as new ligands for positron emission tomography (PET). Such ligands allow the presymptomatic detection of α-synuclein assemblies in brain tissues, which is crucial to early intervention. The results can also be used to develop better model systems of disease and will help to develop methods for producing α-synuclein filaments with structures identical to those in human brains. In the long term, this work could lead to the development of new therapies for Parkinson’s disease. Yang, Y. et al. DOI: 10.1038/s41586-022- 05319-3 Understanding a protein that fuels bowel cancer Tankyrase is an important protein that regulates a wide range of processes relevant to cancer and other conditions, such as diabetes, neurodegeneration and fibrosis. It supports ‘Wnt signalling’, essential for cell division and development and maintaining stem cells. Therefore, tankyrase has received substantial attention as a potential drug target. The protein remains poorly understood, with scientists unsure of how the protein is switched on, how it functions and how to block it without causing unwanted side effects. It self-assembles to form filamentous polymers, but how polymerisation contributes to its function and catalytic activity was unknown. Scientists at The Institute of Cancer Research used cryo-EM at eBIC to investigate the architecture of filaments. In particular, they were keen to identify any potential contacts made by the catalytic domains, as these interactions may control the effect of polymerisation on tankyrase’s activity. They revealed the architecture of a tankyrase filament to be a double helix. Their results revealed extensive interactions between different domains of tankyrase, including those involving the catalytic domain. Based on subsequent biophysical, biochemical and cell-based studies, the researchers proposed that a polymerisation induced allosteric switch regulates tankyrase’s catalytic functions. The scientists suggest tankyrase works by being recruited to a specific site and‘self-assembling’, activating itself by clustering and changing its 3D structure. This work provides novel insights into fundamental biological mechanisms but should also enable the development of novel tankyrase inhibitors and overcome the limitations of currently available molecules. Pillay, N. et al. DOI: 10.1038/s41586-022- 05449-8