Periodic Reporting for period 4 - RELYUBL (Regulation of lymphocyte biology by ubiquitin and ubiquitin like modifiers)
Periodo di rendicontazione: 2020-12-01 al 2021-12-31
A key area of our research explored a poorly understood UBL called UFM1. UFM1 has a similar beta-grasp fold as ubiquitin and has many parallels to ubiquitylation. However, the cellular targets and functions are not understood. Using in vitro reconstitution approaches, biochemistry, structural biology and proteomics approaches we have defined how UFM1 regulates T cell function and immune responses. Our reconstitution approaches have defined the minimal requirements for UFMylation and identified the proteins UFL1 and UFBP1 to together form a functional E3 ligase complex. Further, we identify a scaffold protein CDK5RAP3 that binds to and functions as a substrate adaptor to direct the activity of the ligase complex to endoplasmic reticulum associated ribosomes. As Ufm1 knockout mice are embryonic lethal, we have developed a model where UFM1 is deleted only in T cells or B cells. This reveals that mice lacking Ufm1 exhibit defective immune responses to infection. Applying the tools developed in Aims 1 and 2 together with proteomics and cell signalling analyses we have defined roles for UFMylation in supporting the increased secretory protein biogenesis in activated T cells. Further, we also identified how UFMylation is regulated by the two proteases UFSP1 and UFSP2. The exciting results and novel paradigms have been disseminated by peer-reviewed publications, preprints and presentations at national and international conferences.