The Relationship between CO2 and Ubiquitin in the NF-κB Signalling Pathway

Abstract

Carbon dioxide (CO2) forms a significant proportion of the earth’s atmosphere and plays a crucial role in a number of biological processes including photosynthesis, respiration and acid-base homeostasis. While CO2 is generally unreactive, at physiological temperatures and pressures it is capable of combining rapidly with neutral amines to form carbamates. Moreover, protein carbamylation has been implicated in the induction of subtle conformational changes, eliciting effects within cellular systems and thus functioning as a post-translational modification (PTM). While the labile and readily-reversible nature of this reaction has hindered the study of carbamylation modifications, technological developments continue to facilitate investigations. The ability to chemically trap carbamates under physiologically relevant conditions, for example, has resulted in the identification of ubiquitin as a candidate for modification by carbamylation.

The ubiquitin system has been implicated in a range of biological processes, from gene expression to mediating the stress response. However, it has been demonstrated to play a particularly significant role in the selective degradation of intracellular proteins. Moreover, the ubiquitin-dependent degradation of proteins is involved in the regulation of numerous biological processes, notably the NF-κB signalling pathway. Canonical activation of NF-κB signalling has been shown to be regulated by the K48-linked polyubiquitination-dependent degradation of IκB, a family of inhibitory proteins.

This thesis provides insight into the relationship between CO2 and ubiquitin in NF-κB signalling. Under physiologically high concentrations of CO2, inhibition of K48-linked polyubiquitination was observed, in addition to a reduction in the degradation of IκBα. Furthermore, use of a transcriptional reporter cell line demonstrated a CO2-mediated suppression of NF-κB signalling, in concordance with existing reports. Transfecting cells with a K48R mutant demonstrated the substantial role had by the K48 residue in the regulation of NF-κB signalling, as hypothesised. Overall, this thesis offers a more comprehensive understanding of the ubiquitin-dependent regulation of NF-κB signalling, and the sensitivity of this mechanism to CO2 and carbamylation.