Regulation of Contraction by the thick filaments in skeletal muscle

Muscles make us mobile, and mobility is a major factor determining our quality of life. Unfortunately our muscles get weaker as we get older, and everyday tasks like walking, climbing stairs or even getting up out of a chair get more difficult. Many diseases also lead to more severe forms of muscle weakness, and some of these affect young children. In general there is no effective treatment for muscle weakness. Using muscles- exercising them- does make them stronger, but this is not always possible, particularly for those suffering from diseases that affect the muscles. This proposal follows up some new discoveries about how the strength of healthy muscles is controlled. It aims to understand those new control mechanisms better so that it will be possible to design and test potential drugs to intervene in muscle control and boost the strength of weak muscles.

Muscles are built from long strings of a basic microscopic building block called a sarcomere. Each sarcomere contains an array of two types of filament, one wider than the other, and these are called thick and thin filaments. The relative sliding of these two sets of filaments is responsible for muscle shortening. When a signal from the brain travels along a nerve and reaches a muscle, it triggers release of calcium from stores inside the muscle cell. The calcium binds to the thin filaments, causing a change in their structure that allows the filaments to slide, and the muscle contracts. This change in thin filament structure is quite well understood, but it works like an OFF/ON switch- it controls when the muscle contracts, but not how strongly. Recently another type of muscle control was discovered that works by changing the structure of the thick filaments. These changes in thick filament structure control the strength and speed of muscle activation, and how fast they relax, but we still know very little about how they work. Thick filament structure also controls how much energy a muscle uses when it is resting. Since about a third of the weight of our bodies is muscle, understanding how the OFF state of muscle is controlled might help to combat some very different health problems, by allowing muscles to be used to burn off unwanted calories.

At present we don't know how these OFF and ON states of muscle are controlled- we don't know how the structure of the thick filament changes, and what controls those changes. In this project we will apply the technical advances provided by two of the brightest X-ray sources available worldwide (in France and the USA) and a fluorescence based method that we developed to tag the thick filament proteins to measure the changes in thick filament structure when isolated muscle cells contract. These methods will allow us to answer the key questions about how the structure of the thick filaments controls the strength and speed of muscle contraction and, no less important for the way muscles are used in the body, how the speed of muscle relaxation is controlled.

The answers to these questions will allow us to develop a detailed picture of how the OFF and ON states are controlled by changes in thick filament structure in healthy muscle. This in turn will enable us to suggest ways in which these states might be controlled by drugs, and to develop ways to assess the value of potential new drugs. Finally, since very similar changes in thick filament structure occur in heart muscle using the same protein components, we expect that the results of this project will also be useful in guiding an analogous approach to controlling the strength of heart muscle, and therefore in developing potential new treatments for heart disease.

Contraction of skeletal muscle is usually considered to be regulated by the intracellular calcium concentration, mediated by structural changes in the actin-containing thin filaments. However it is becoming increasingly clear that the strength, speed and metabolic cost of contraction, and the speed and timing of relaxation, are largely controlled by a separate set of regulatory mechanisms involving the myosin-containing thick filaments. Those mechanisms are poorly understood at present, although they offer a powerful new approach to the design of molecular therapies for muscle disease and weakness. The project will apply state-of-the-art time-resolved synchrotron X-ray and polarized fluorescence techniques to elucidate those mechanisms at the molecular level in the intact filament lattice of intact and demembranated cells from mammalian skeletal muscle. A key aim is to determine how thick filaments are activated following the intracellular calcium transient. Two mechanisms seem most likely: direct mechanical activation of thick filaments and inter-filament signalling by myosin binding protein-C (MyBP-C). We will determine the roles of those two mechanisms in thick filament activation, and clarify the wider role of MyBP-C in modulating muscle performance. In an analogous approach we will determine the roles of load, filament sliding and inter-filament signaling on the kinetics of muscle relaxation, and elucidate the underlying molecular mechanisms in the thick and thin filaments. Finally we will determine the structural basis of the enhancement of muscle performance following recent activity and the role of phosphorylation of the myosin regulatory light chain in that process. The results of these studies will facilitate the development of molecular targets and assays for potential therapeutic modulation of contraction in diseases of both skeletal and heart muscle.

Who will benefit from this research, and how?

1. The elderly. There are more than 10 million people over normal retirement age in the UK, and worldwide there are more than 600 million people aged over 60, many of whose lives are affected by weakness of their skeletal muscles, as a normal part of the ageing process. This research aims to open up new approaches to the treatment of muscle weakness through a better understanding of the normal mechanisms by which muscle strength is controlled.

2. Those suffering from muscle weakness due to disease. This would include patients suffering from congenital and adult-onset neuromuscular and neurodegenerative disorders, obesity and its associated disorders, and cachexia. Again the potential benefits would come from new approaches to the treatment of muscle weakness through a better understanding of the normal mechanisms by which muscle strength is controlled.

3. Those suffering from heart failure. Over three-quarters of a million people in the UK alone suffer from heart failure, and this is another important factor influencing quality of life for many older people. Because of the close similarity between the regulatory pathways under study in this proposal in skeletal and heart muscle, we expect that the results of the proposal will guide the development and testing of potential new drugs that improve contractility in the heart, as a means to counteract heart failure.

4. Those suffering from diabetes and obesity. More than three million people have been diagnosed with diabetes in the UK alone, and worldwide over 400 million people were obese in 2008. The potential impact of the present proposal for this group relates to the dominant impact of skeletal muscle metabolism on blood glucose levels and body mass, and the possibility of long-term control of these factors by modulation of the rate of ATP utilization by resting muscle.

5. Commercial organisations. The long-term beneficiaries within the private sector are likely to be companies involved in the further development of small-molecule therapeutics arising from the results of the proposal. We have held a series of meetings with a company specializing in small molecules that modulate the contractility of skeletal and heart muscle, as described in Pathways to Impact, and this would be our preferred initial route for exploring the commercialization of potential new targets arising from the present proposal.

6. Academic beneficiaries. These include researchers in muscle physiology, muscle development, sports and exercise physiology, ageing, skeletal muscle disease, heart disease and obesity, and clinicians working in the corresponding specialities, as described in the section Academic beneficiaries.

7. Staff employed on the project. The employed staff will benefit from enhanced research, technical and planning skills. The project will be a key step enabling the PDRA to be ready to undertake an independent research career. More widely, the project will help to maintain the shrinking pool of researchers with the necessary skill-base to perform demanding holistic or systems-level approaches to elucidate physiological mechanisms in intact muscle.

Most of the above potential benefits are long-term, and the likelihood of their realization would be assessed during the project in order to prioritise future work between the potential targets listed above.