In accordance with the proposals outlined in fellowship proposal, we successfully designed and performed screens for novel interacting proteins and potential substrates for the rhomboid proteases. This took the form of mass spectrometry-based screens and novel cell biological assays designed in-house. Furthermore, we have generated loss-of-function model systems for the analysis of the hits from the screen, which are currently being used to validate the putative substrates identified by the screen. This research has been communicated at various regional and international meetings and conferences. Overall, the main objective of the project has been fully met and the results are likely to be the foundation for further in-depth characterisation and subsequent publications.
During the period of the fellowship funding, a review on the rhomboid family of intramembrane proteases has been published in an open-access peer-reviewed journal, which has helped to increase awareness of the significance of fundamental research on rhomboid proteases, and the importance of substrate discovery:
Lastun, V.L. Grieve, A.G. & Freeman, M. (2016) Substrates and physiological functions of secretase rhomboid proteases. Semin Cell Dev Biol, doi: 10.1016/j.semcdb.2016.07.033
Further to this, a methodology paper is currently in press, describing cell biological assays for rhomboid protease function. This will help others in the field to efficiently screen for rhomboid protease substrates:
Moncada-Pazos, A. & Grieve, A.G. (2017) A simple cell-based assay for the detection of surface protein shedding by rhomboid proteases. Methods in Molecular Biology, In Press.
Outside of the scope of our initial plan, we investigated the function of rhomboid family member 2, RHBDF2 (also known as iRhom2), and its role in inflammation through regulation of the metalloprotease, TACE. This work represents the discovery of key mechanisms that control the cleavage and release of the primary inflammatory cytokine, TNF. This research on RHBDF2 and inflammation has been published in an internationally recognised open-access journal, and widely communicated through social networking:
Grieve, A.G. et al. (2017) Phosphorylation of iRhom2 at the plasma membrane controls mammalian TACE-dependent inflammatory and growth factor signalling. Elife, 6, doi: 10.7554/eLife.23968.
In addition to many talks and conferences where this research has been presented, we wrote a description of our work in the journal using lay terms, so that the impact of fundamental biological research is communicated as widely as possible (known as an ‘eLife digest’). The aim of which was to narrow the gap between active scientists and the general public with particular interest in the biosciences.
