Transcriptional and Post-translational Regulation of Cytosolic Carbonic Anhydrase in Rainbow Trout (Oncorhynchus mykiss) and Zebrafish (Danio rerio)

Abstract

The enzyme carbonic anhydrase (CA) contributes to multiple physiological processes by catalysing the reversible hydration of carbon dioxide. However, regulation of CA activity in response to homeostatic challenges remains poorly understood. The objectives of this thesis were to investigate whether CA is transcriptionally regulated by cortisol in fish and whether post-translational modification (PTM) of CA occurs in fish. The results of an in vivo reporter assay used to investigate potential transcriptional regulation of zebrafish, Danio rerio, cytoplasmic CA (CAc) were inconsistent, and it remains unclear whether zebrafish CAc is regulated transcriptionally by cortisol. Phosphorylation of rainbow trout, Oncorhynchus mykiss, CAc was predicted from in silico analysis of the putative amino acid sequence and confirmed by Western analysis of phosphoprotein levels following in vitro incubation of CA, purified from trout gill, under conditions designed to potentiate endogenous kinases. Again using in vitro incubations designed to potentiate endogenous kinases and phosphatases, changes to the phosphorylation state of CAc were found to modulate its enzymatic properties. These findings suggest that CA activity may be regulated by signalling pathways that activate cellular protein kinases, and future work should focus on identifying these pathways.