Determining how mutations in actin lead to skeletal muscle weakness

Skeletal muscles are composed of a large number of cells called myofibres. Each myofibre have myofibrils consisting of sarcomeres in series and parallel. Each sarcomere is formed by overlapping arrays of thick filaments and thin filaments also known as myosin and actin filaments. Myosin and actin filaments represent the contractile machinery.

There are many acquired and inherited diseases affecting myosin and actin filaments. Among them, actinopathies are the most common and are caused by mutations in actin. Actinopathies are usually life threatening, as they are associated with severe weakness affecting limbs and respiratory muscles. How mutations in actin lead to muscle weakness remains obscure and will be addressed in the present research project. Deciphering this will undoubtedly help designing efficient therapeutic interventions.

Importance and main objective: Diseases related to mutations in actin are characterized by severe skeletal muscle weakness often leading to premature death. The understanding of this group of disorders has advanced in recent years through the identification of the causative ACTA1 gene mutations. Nevertheless, the exact mechanisms leading to muscle dysfunction remain unclear, which hampers the development of efficient therapies. Therefore, the main objective of the present research project is to clearly characterise the cascade of molecular and cellular events triggering muscle wasting.

Methods: We will mainly use muscle samples and isolated myofibres from two transgenic mouse models perfectly recapitulating the human condition and expressing either His40Tyr or Asp286Gly single amino acid substitution in actin. We will then perform a broad range of experiments including small-angle X-ray scattering, polarised fluorescence, myofibre mechanics, confocal microscopy, immunohistochemistry and micro-array analysis.

Feasibility and outcomes: The availability and knowledge of these unique techniques in combination with experience in undertaking challenging studies put us in a unique position to tackle the problem posed. We expect to demonstrate that mutations in actin directly impair actin filament extensibility, inducing a disruption (i) of myosin binding and (ii) of the intrinsic force-generating capacity, directly promoting myofibre atrophy. Successfully proving this would grandly facilitate the design of efficient therapeutic interventions that would target the actin-myosin interface.

1. Impact by training:
As with any reputable scientific centre we are concerned that young scientists who come for training should be given as wide a scope as possible in order to keep their future scientific options open. This project will allow the development of the appointed individual to develop a range of unique and specialist skills such as:
(i) Learning new techniques; and
(ii) Gaining communication skills by presenting his/ her work to the scientific and medical communities and to a more general audience.

2. Impact on basic research and academia:
The effort to tackle rare diseases is hampered due to the complex phenotypes and low number of specialized research groups. The present research project combining state-of-the-art methods and unique techniques has the potential to move the field into completely uncharted scientific territory. We predict that the findings of the present proposal, at the interface between physiology and biophysics, will be of great interest for:
(i) Geneticists who focus on gene mutations and their consequences;
(ii) Structural and cell/ molecular biologists, who are interested in actin structure and function;
(iii) Biophysicists who use similar techniques including small-angle x-ray scattering;
(iv) Physiologists who investigate skeletal/ cardiac muscle structure and function in health and disease; and
(v) Clinicians who try to understand the pathophysiology of muscle diseases.

3. Impact on the clinic:
Rare diseases affect more than 25 million people in Europe. Only a few of these disorders are well enough understood to be treated effectively. Thus, rare diseases are still an important public health challenge. Our research project has the potential to contribute to the nation's health by
(i) Generating attention from affected patients, family members, general practitioners, myologists, neurologists, pathologists and physiotherapists; and
(ii) Setting an example for tackling other rare (muscle) diseases such as cardiomyopathies, which are also frequently related to mutations in genes encoding actin.

4. Impact on pharmaceutical industry:
Actinopathies start at a very young age. Hence, the economic consequences are large (loss of ability to work, large medical expenses) and the impact on family members is enormous, as they need to take care of the affected patients as long as they live. The fact that we will unravel the pathophysiological mechanisms underlying weakness in a group of rare disorders termed actinopathies will:
(i) Provide a clear understanding of how mutations in actin cause disease; and
(ii) Give opportunities for industrial applications; in other words, help to increase the speed of developing therapeutic interventions by identifying potential drug targets.

5. Impact on society via engagement of the public in science:
The PI of this application is a member of various scientific societies and takes active roles in trying to engage interested members of the public, particularly younger people, in science. The Centre of Human and Aerospace Physiological Sciences has an outstanding track record in linking its research to public engagement and activities for engaging young people. This includes endeavours such as "Mission Discovery", a weeklong summer school using the physiology of human space flight to facilitate and engage children in biomedical science and designing experiments, which run on the International Space Station:
http://www.kcl.ac.uk/newsevents/news/newsrecords/2014/January/missiondiscovery-lift-off.aspx)