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Structural and functional characterisation of the novel endoribonuclease SLFN14

Secondary Supervisor(s): Dr Alexandre Slater

University of Registration: University of Birmingham

BBSRC Research Themes:

• Understanding the Rules of Life (Structural Biology, Systems Biology)

Project Outline

The novel hematopoietic-specific SLFN14 endoribonuclease is responsible for general translation control where human SLFN14 missense mutations cause an inherited platelet-based bleeding disorder with significant platelet dysfunction, but the precise mechanism of disease is unknown. To investigate the role of SLFN14 in platelet biogenesis, we will employ biochemical, molecular and cellular biology techniques, and structural analysis via X-ray crystallography and cryo-electron microscopy (cryo-EM). Employment of selective inhibitors for the major degradation systems in platelets will reveal the degradation pathway underlying the SLFN14 function/dysfunction. Binding partners of SLFN14 in platelets will be fully characterised using co-IP assays in WT samples followed by mass spectrometry analysis of SLFN14 containing complexes. Novel nanobodies will be raised against SLFN14 and characterised as potential thrombotic targets. We will interrogate the protein structure of wild type SLFN14 and the effect of SLFN14 mutations which can then be selectively targeted therapeutically in patients but also advance our understanding of the role of SLFN14 in MK maturation, platelet biogenesis and ribosomal elimination in newly produced platelets. Overall, this work will provide novel insights into mechanisms underlying SLFN14 and other bleeding disorders, while also identifying pathways regulating platelet function that may be novel targets for the development of anti-thrombotic therapy.