Yeast models to study mutations in SURF1 and MPV17 involved in human mitochondrial disorders

by Robert Reinhold
Doctoral thesis
Date of Examination:2011-11-25
Date of issue:2011-12-06
Advisor:Prof. Dr. Peter Rehling
Referee:Prof. Dr. Peter Rehling
Referee:Prof. Dr. Mikael Simons
Persistent Address: http://dx.doi.org/10.53846/goediss-3553

 

 

Files in this item

Name:reinhold.pdf
Size:17.1Mb
Format:PDF

The following license files are associated with this item:

Abstract

English

Mitochondrial diseases are fatal disorders mainly affecting highly energy dependent tissues such as brain, heart, liver and muscle. These severe disorders can be caused by mutations affecting mitochondrial- or nuclear-encoded proteins. The molecular function of many nuclear-encoded mitochondrial proteins involved in disease is unknown so far and therefore, the mechanisms that lead to disease are enigmatic. In this study, point mutations in SURF1 and MPV17 were analyzed that were found in patients of the Leigh Syndrome and the Mitochondrial DNA Depletion Syndrome respectively. These missense mutations affect highly conserved and therefore potentially functional important amino acids. The function of SURF1, a highly conserved early assembly factor of the cytochrome c oxidase, has been studied using its yeast homolog Shy1. Shy1 is involved in the maturation and translational feed back regulation of Cox1, the central subunit of the cytochrome c oxidase. In this study, it was shown that patient point mutations affecting G124 in SURF1 lead to rapid degradation of the protein after mitochondrial import. In contrast, patient mutations that lead to an Y274D exchange in SURF1 do not alter mitochondrial import or stability but showed increased association to a newly identified cytochrome c oxidase assembly intermediate. Hence, SURF1Y274D shows impaired function. Using the corresponding yeast Shy1Y344D as a model, dual functionality of SURF1/Shy1 could be shown. Shy1Y344D releases cytochrome c oxidase assembly intermediates from translational feed back regulation of Cox1 but fails to promote cytochrome c oxidase assembly at later steps. Mutations affecting MPV17, a mitochondrial inner membrane protein of previously unknown function, lead to loss of mitochondrial DNA. In this study Sym1, the yeast homolog of MPV17, was used as a model to study the biogenesis and function of MPV17/Sym1. It could be shown that Sym1 forms high molecular weight complexes of 120 and 220 kDa in the mitochondrial inner membrane. In contrast to most polytopic inner membrane proteins, Sym1 is imported in a TIM23 dependent manner into mitochondria. The function of Sym1 was analyzed by electrophysiological characterization of Sym1, isolated from yeast mitochondria. Sym1 showed channel activity, which indicates a pore forming function of Sym1/MPV17 in the mitochondrial inner membrane.
Keywords: SURF1; MPV17; mitochondrial disorder
 
Statistik