Actin Aggregation Leads to Mitochondrial Dysfunction and Ageing

Actin is an essential protein, which is involved in many of the important functions of a cell, from structural integrity, to how it moves and senses its environment. The role of actin in these events is enabled by its dynamic nature, as it can be readily assembled into highly regulated filaments and disassembled again as required. Under certain circumstances this process goes wrong and instead of forming dynamic filaments, actin aggregates into large clumps. Such clumps are found in the neurones of our brains as we age and are prevalent in patients suffering from neurodegenerative disorders. It is likely that actin aggregates contribute to the processes of ageing and neurodegeneration, but little progress has been made in establishing whether this is the case. Budding yeast have been successfully used to investigate how cells function for many years. I intend to use yeast to study how and why actin aggregates form, and to establish what effects they have on a cells health. I will also examine how actin aggregation affects the rate at which human cells, grown in the laboratory, age. These approaches will advance our understanding of age related diseases, such as Alzheimer‘s, and facilitate the development of therapies.

It has recently been identified that actin has a conserved role in the processes of ageing and apoptosis. The aggregation of actin filaments have been implicated in neurodegeneration. In fact a particular actin aggregation, the actin/cofilin rod (ACR), promotes amyloid plaque formation, and so is likely to participate in the progression of Alzheimer‘s disease. However the mechanisms through which actin aggregates form and how they exert toxicity are largely unknown. My previous studies have identified a number of conditions under which the model eukaryotic yeast Saccharomyces cerevisiae forms actin aggregates. Their formation consistently leads to mitochondrial dysfunction, the accumulation of reactive oxygen species (ROS), premature ageing and apoptosis. As the mechanisms of actin regulation are also highly conserved, budding yeast are an attractive model system in which to study the toxic effects of actin aggregation. Actin aggregation leads to the constitutive activation of Ras signalling, which perturbs mitochondrial function leading to ROS generation. I will investigate the nature of this activation and identify the targets of the actin aggregate induced Ras signalling cascade. The composition of actin aggregates is likely to be central to their toxic effects, I will therefore purify aggregates from yeast and identify the protein content using a proteomic approach. My preliminary investigations have also established that ROS accumulation occurs in actin aggregated cells as a result of a defective mitochondrial electron transport chain (ETC). The effects of aggregates formation on ETC composition and respiratory function will therefore be investigated. It is essential that the proteins, which modulate actin aggregate formation, are identified and characterised. My preliminary studies have identified Smy1p as an inhibitor of aggregate formation. Smy1p belongs to the type-1 kinesin family and has the potential to link mitochondrial regulation with the regulation of actin dyanmics. I will therefore characterise the effects of Smy1p on mitochondrial function and actin regulation. As an extension of Preliminary data observed I will conduct pioneering studies into the effects of actin aggregation on ageing in a human cell culture system. These experiments will examine the effects of ACR formation and persistence in a human lung fibroblast cell line on ageing and senescence. The information, which will be obtained from these experiments, will greatly advance our knowledge of how actin aggregation imparts toxicity. This has significant medical implications and will facilitate the development of therapeutic strategies to combat a variety of age-related diseases.