Thiol-disulfide redox switches in protein kinases and their role in cardiovascular health and disease

Whilst oxidants may contribute to the pathogenesis of cardiovascular disease, many of them also function as important signalling molecules, carrying vital information about cellular redox state. Oxidants are crucial for adaptation of the cardiovascular system during health, as well as during disease. Oxidants can regulate biological processes by inducing structural modifications of proteins, which can changes their function. This application seeks funds to capitalise on our recent identification of new redox sensors. Our aim is to define the role of these newly identified redox sensors proteins in cardiovascular health and disease. These studies may provide new insight to targets for drug development that improve human health in the setting cardiovascular disease. This is important as cardiovascular disease is the leading cause of death in the world, including in the United Kingdom.

We have previously discovered several proteins in the cardiovascular system that are regulated by changes in their oxidation state. These proteins play crucially important roles in the healthy function of blood vessels and the heart. However, these redox-regulated proteins can also become aberrantly stimulated during stress scenarios associated with increased oxidant levels (oxidative stress), events that can contribute causatively to disease pathogenesis. At the moment our understanding of how changes in oxidative stress regulate proteins in the cardiovascular system is far from complete, an issue that this application seeks to address. We have recently identified a host of proteins in the heart that were previously not known to be modified by oxidants, and our objective now is studying them to understand the impact this have on their enzymatic activities, as well for cardiovascular function. In addition to studying these proteins at the biochemical and cell biological level, we also have generated novel transgenic mouse models in which the redox sensing has been replaced so they cannot sense or transduce oxidant signals. By studying these redox dead mice, or tissues or cells isolated form them, and comparing their response to wild type controls, we will be able to define the roles these protein sensors play in the maintenance of cardiovascular health, and how they may be dysregulated during disease. To achieve our aims we will use a multidisciplinary approaches, with in vivo cardiovascular phenotyping (telemetric blood pressure, echocardiography, Pressure-Volume analysis) together with ex vivo functional tissue studies (isolated hearts, isolated blood vessels) and biochemical analyses (Western immunoblotting, kinase activity assays, histology). Protocol with mice will be randomised and blinded analysis and subject to full power analysis to ensure adequate sample size is utilised. Ultimately, these studies may provide important clues about new drug targets.

This application focuses on the functional impact of protein kinase oxidation and its potential role in the pathogenesis of cardiovascular disease. RSK1 and PKG Ialpha are intensely studied kinases, and our identification of novel mechanisms involved in its regulation may help generate new ideas in many research areas. These kinases, along with NIMA-related kinase 7 (Nek7), ribosomal protein S6 Kinase Beta 2, also known as p70S6K2 (S6K2) are expressed in multiple tissues and has a broad impact on many cellular functions. Consistent with this, dysregulation of kinase-dependent signalling is widely implicated in the pathogenesis of multiple diseases, not just those of the cardiovascular system - but also cancer and neurological disorders. Our pilot data clearly indicate that RSK1 oxidation to an intradisulfide occurs during scenarios relevant to myocardial disease, with some early preliminary indication this is also the case for PKG Ialpha oxidation to the intrdiculfe. A wide range of scientists in different albeit complementary specialties will potentially benefit from the proposed studies, including physiologists, biochemists, structural biologists, medicinal chemists and pharmacologists. Given the ultimate translational potential of these studies, the project could impact positively on healthcare professionals at multiple levels within this sector. Ultimately, new therapies could arise from this work, which would aid clinicians and their colleagues in reducing the burden of highly prevalent cardiovascular diseases such as hypertension heart failure. Effective therapies against these highly prevalent conditions that negatively influence the health and wealth of society would strengthen the position of policy makers, healthcare managers, doctors and related staff and end-users as it could save an enormous amount of resource that could then be reallocated to address other problems. As kinase dysregulation is also implicated in the pathogenesis of cancer and some neurological disorders, our proposed studies may potentially lead to new understanding or therapies that alleviate those conditions. This would provide further significant wealth benefits to society, as well as reducing other negative aspects associated with the high morbidity and mortality caused by those diseases. Overall, it is evident that society at many levels may have much to gain from a better understanding of the redox control of protein kinases. This potentially could have a huge impact - not least because of the translational potential that could help in the fight against several diseases highly prevalent in the aging population.