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Title: Novel biomarkers for arthritis : the role of P2X7 receptor in transglutaminase 2 export and activation
Author: Griffiths, Rhiannon
Awarding Body: Cardiff University
Current Institution: Cardiff University
Date of Award: 2017
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Transglutaminase 2 (TG2)-mediated stabilization of extracellular protein assemblies has a pivotal function in tissue repair. However, aberrant TG2 activity has been linked to fibrosis and autoimmunity. There is also evidence that post-translational modification of proteins can occur in a manner that is specific to disease, for example citrullinated peptides seen in rheumatoid arthritis. TG2 modifies proteins in several ways, including through transamidation, esterification and deamidation of target glutamine residues. There is evidence of increased expression and activity of TG2 in osteoarthritis, potentially leading to the generation of protein modifications, which could act as biological markers of disease. The mechanism of TG2 release by cells controls its extracellular activity is unconventional and enigmatic. Our group has shown that TG2 export is linked to purinergic signalling, and implicated P2X7 receptor activation. P2X7R has several activation states; ATP stimulation causes ion channel opening,allowing membrane depolarization and Ca2+ entry into the cell. Prolonged stimulation leads to ‘large membrane pore’ activity, however the identity of this pore is unknown as there is conflicting evidence suggesting either dilation of the P2X7R channel itself or an interaction with an alternative plasma membrane channel. The aim of this thesis is to elucidate mechanistically the process by which cells export TG2 and control its activation, alongside investigating the P2X7R pore. We have also looked at the involvement of GTP regulation of TG2 activity in its secretion and whether the TG2 activator, thioredoxin, can be secreted via the same pathway. Through the use of pharmacological agents, we have shown that pannexin-1, a plasma membrane hemichannel proposed to interact with P2X7R, is unlikely to be the pore-forming component of P2X7R activation. A gain of function mutation in P2X7R expressed in HEK293 cells shows enhanced TG2 externalization from cells, correlating with increased pore activity,implicating P2X7R itself. Thioredoxin, a TG2 activator, is co-secreted. To confirm the transferability of our findings in this cell model to the innate immune response, human peripheral blood monocytes were differentiated into M1 macrophages and P2X7R mediated TG2 export assessed. Investigating this process will unravel a novel secretory pathway potentially used by select proteins, including potent signals regulating inflammation.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available