Investigating the role of AURKAIP1 in mitochondrial gene expression

Abstract

Mitochondria contain their own genome (mtDNA), which encodes 13 polypeptides essential for oxidative phosphorylation. Dysfunctional mitochondria have been associated with disease and the ageing process. Expression of mtDNA requires import of many nuclear-encoded polypeptides that are synthesised in the cytosol and imported into the mitochondrion. As many as 300 nuclear-encoded mitochondrial proteins are estimated to be unidentified or their functions remain unknown. The importance of understanding the nuances of human mitochondrial gene expression machinery has been highlighted by the predisposition of paediatric mitochondrial patients with nuclear gene mutations that manifest as mitochondrial translation defects. The aim of this research was to further the understanding of mitochondrial gene expression by characterising the protein AURKAIP1, which was identified during a screen of 224 candidate proteins as being likely to have a role in mitochondrial gene expression. By mitochondrial sub-fractionation techniques I was able to show that AURKAIP1 is a mitochondrial matrix protein. I have shown that AURKAIP1 is involved in mitochondrial gene expression since depletion of AURKAIP1 severely depletes the levels of mitochondrial-encoded proteins ND1, COXI and COXII. In addition, the overexpression of AURKAIP1 in mammalian cell lines causes a dramatic decrease in the de novo synthesis of mitochondrial polypeptides, which was studied by labelling nascent peptides with [35S] labelled methionine. The overexpression of AURKAIP1 also reduces the steady state levels of many mitoribosomal proteins, suggesting the reduced level of mitochondrial gene expression is due to a reduction in the number of functional mitoribosomes. I have also demonstrated a novel interaction between AURKAIP1 and the mitochondrial protein, p32, which is essential for mitochondrial translation. A paediatric patient with severe mitochondrial disease was found to have mutations in p32, which highlights the importance of mitochondrial gene expression and the need for further study into the factors involved.