Role of mitochondrial Complex I in cellular response to hypoxia

Where there?s life, there is energy. In our body energy is generated by mitochondria often referred as power stations of the cell. Most of the oxygen from the air we breathe is consumed by mitochondria to produce energy during the process called cellular respiration. This energy is used for many needs of the body such as heart beating, brain activity, kidney functioning as well as synthesis of proteins and DNA inside all our cells. Hypoxia is a lack of oxygen in the body or any organ and can damage cells and tissues. Hypoxia can be caused by many factors, including disease, pathology or trauma and it also occurs during heart surgery. Usually, hypoxia is followed by restoration of oxygen supply (reoxygenation), which makes the damage greater and in some cases irreversible. Mitochondria are responsible for that damage, when they form toxic agents known as free radicals. Enzyme called Complex I is the most important and least understood key component of the mitochondria and it initiates the processes of energy production during respiration of the cell. We have recently found that Complex I is very important in hypoxia/reoxygenation and can be damaged in that process. Its damage results in lack of energy in a cell, generation of harmful free radicals, oxidative stress and eventually lead to tissue injury.

We are going to investigate the involvement of mitochondria in the processes of hypoxia/reoxygenation by our novel biochemical approach and by advanced non-invasive cellular respirometry techniques for measuring how cells consume oxygen. Also we will be using fluorescent confocal microscopy, special technique to look directly at the different parameters in a single alive cell. Our project will help to develop drugs that can selectively modulate the activity of Complex I in order to lessen tissue damage in brain and heart disorders. In addition it would help us to develop a strategy for early diagnostic and prevention therapy for initial stages of neurodegenerative disorders such as Parkinson?s and specific heritable diseases. Our research will help to develop new drugs that may stop or slow down the death of nerve cells. In addition, knowledge on how hypoxic signals are carried out inside the cell will allow clinicians to suppress growth and invasion of cancer which is often associated with lack of oxygen in the tumour.

Mitochondrial complex I plays a critical role in regulating energy generation in cell and it is involved in a number of clinical conditions such as neurodegeneration (Parkinson?s disease, Leber?s optic neuropathy), muscular pathologies and processes of ageing. At the same time ischaemia/reperfusion injury mediated by mitochondria is a major cause of heart diseases, as well as stroke; these two are the leading causes of death in the UK, ranking before cancer and cause more than 30% of deaths. This proposal is aimed at unraveling the role of mitochondrial complex I in such pathological conditions translating new findings to clinical medicine.

Complex I has a high control over the respiration rate and therefore it is likely that the moderate changes in its activity would make a strong effects on the overall rate of oxidative phosphorylation. In addition Complex I is also a source of detrimental free radicals, as well as a target for oxidative damage by them. I intend to clarify the role of Complex I in cellular response to hypoxia and identify possible key points of control of the enzyme my physiological and pharmacological modulators. I propose to investigate Complex I behaviour in detail using classical biochemical as well as novel non-invasive cell-imaging approaches.

Firstly, I will identify structural differences between two Complex I states in isolated mitochondrial membranes from brain and heart, characterise them by cross-linking and amino acid specific covalent reagents. Further I will study Complex I activity and factors influencing it in several cultured cell lines including myocytes and neurons during processes of hypoxia and reoxigenation. I plan to use biochemical and live cell imaging techniques to carry out functional studies of effect of pharmacological agents on Complex I and how they influence cell viability in hypoxia/reoxygenation. The proposed line of research will clarify the physiological and pathophysiological consequences of complex I activity changes and ROS production during hypoxia/reoxygenation and nitric oxide exposure.

Prolonged hypoxia initiates a sequence of cellular responses via the hypoxia-inducible transcription factors (HIFs). It has been proposed that mitochondrial ROS, oxygen redistribution and other transcriptional pathways are involved in HIF-1alpha stabilisation at low oxygen concentrations, but the relative contribution of these mechanisms in hypoxia is not clear. I will therefore also study the involvement of cellular components in accumulation of HIF-1alpha via ROS-dependent and independent mechanisms.

In the proposed project I am going to bring together two essential aspects of hypoxic studies important for development of cardio- and neuroprotective therapy and anticancer interventions. Understanding how and why Complex I activity is modulated in cells will elucidate the role of mitochondria in ischaemic/reoxygenation injury, muscular- and neurodegeneration as well as age-related loss of muscle function. Such understanding could lead to the development of new clinical diagnostic tools and presents opportunities for therapeutic intervention. At the same time unravelling of involvement of respiratory chain in HIF-1alpha stabilisation will help to design strategies for anticancer treatment.