Periodic Reporting for period 3 - C-CLEAR (Complement: to clear or not to clear) Reporting period: 2021-07-01 to 2022-12-31 Summary of the context and overall objectives of the project Mammalian complement recognizes a variety of cell-surface danger and damage signals to clear invading microbes and injured host cells, while protecting healthy host cells. Improper complement responses contribute to diverse pathologies, ranging from bacterial infections up to paralyzing Guillain-Barré syndrome and schizophrenia. What determines the balance between complement attack reactions and host-cell defense measures and, thus, what drives cell fate is unclear. The project tries to elucidate molecular mechanisms that underlie the balance between complement attack and host-cell defense, which determine cell fate. Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far In the first phase of the project, a series of new experiments have been set up. Significant progress has been made by cryo-EM single-particle analysis and tomography providing new opportunities to reveal key steps in complement activation and defense. Moreover, a new concept is emerging that may indicate a potential mechanism that makes cells more prone to complement attack. Overall the project has had a very good start, but further work is needed and planned to substantiate the stimulating intermediate results.Progress was hampered by the COVID outbreak, causing temporarily a complete shut-down (9 weeks) of all experimental activities not direly related to SARS-2 research. Data analysis was continued, computational projects were started and research with regard to membrane organization was focused on SARS-2. After the (complete) shut-down period research was restarted with restrictions. Overall, all three parts of the project are in progress, yielding new insights and further defining our research approach. Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far) We will combine crystallography, cryo-EM, cryo-ET and high-resolution microscopy to resolve complement complex formations and reactions on membranes.Massive cryo-EM data analysis addressed the inherent flexibility of complexes yielding insights into complement activation (part I); the emergence of the new concept of membrane organization needs further substantiation (part II); and, unexpected aspects of complement activation suggest further optimization of the experimental conditions for homogeneous complex formation (part III). For all parts experimental conditions are well poised for subsequent experiments, addressing hitherto unresolved aspects of complement activation.
