AbstractsBiology & Animal Science

Sarco(endo)plasmic reticulum Ca2+ ATPase (SERCA) sequesters Ca2+ from the cytosol to: i) aid in muscular relaxation and ii) refill SR Ca2+ stores for release upon subsequent stimulus for contraction. SERCA function is modulated by physical interaction with phospholamban (PLN), where phosphorylation of PLN relieves its inhibitory action and increases SERCA activity. Protein-protein interactions have been shown to be protective against heat-induced SERCA2 modifications within HEK-293 cells (Fu & Tupling, 2009). Unpublished work from our lab has specifically shown SERCA2 protection in whole left ventricle (LV) when co-expressed with its regulatory modulatory protein, PLN (Gamu & Tupling, unpublished). The current thesis builds on this work by pairing vasomotor functional data from an isolated artery segment preparation with biochemical data to further explore the protective interaction between PLN and SERCA2. Peroxynitrite (ONOO-) is an oxidant known to decrease SERCA2 function through irreversible damage (Viner, Williams, & Schöneich, 1999). Thoracic aorta from wild type (WT) and phopsholamban null (PLN-/-) mice were exposed to varying concentrations of ONOO- (80 μM, 150 μM and 300 μM). Following this oxidative incubation, vasoactivity was assessed and compared. Furthermore, specific SERCA2 activity and structural adaptations through 3-NY formation were investigated. Interestingly, ONOO- incubation (150 μM) elicited genotype differences throughout the incubation period, where PLN-/- vessels had an increased basal tension response compared to WT vessels (p<0.05). Also, following 150 μM ONOO- incubation, EC50 to SNP-induced relaxation was significantly increased in PLN-/- vessels compared to WT (p<0.05). Both genotypes experienced reductions in PE-induced contractility, highlighting the ability for ONOO- incubation to decrease contractility (p<0.05) though total SERCA2 activity was significantly reduced only following the 300μM ONOO- incubation (p<0.05). No significant differences in SERCA2 activity were found between genotypes, indicating SERCA was not protected through its interaction with PLN from exposure to high ONOO-. This data shows high concentrations of ONOO-incubation is able to significantly reduce SERCA2 activity, however we only have some evidence to support that PLN may be protective. This stands as a basis for future work to be completed regarding the protective nature of PLN and SERCA2 interaction