Publication Date

2012-10-19

Availability

Open access

Embargo Period

2012-10-19

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PHD)

Department

Biochemistry and Molecular Biology (Medicine)

Date of Defense

2012-10-11

First Committee Member

Antonio Barrientos

Second Committee Member

Kenneth E. Rudd

Third Committee Member

Feng Gong

Fourth Committee Member

Miguel A. Perez-Pinzon

Fifth Committee Member

Rosemary A. Stuart

Abstract

Cytochrome c Oxidase (COX) is a multimeric mitochondrial protein complex acting as the terminal enzyme of mitochondrial respiratory chain (MRC). It catalyzes the transfer of electrons from cytochrome c to molecular oxygen that is reduced to water. The efficiency of the electron transfer can be modulated in response to oxygen availability by differential expression and assembly into the holoenzyme of subunit isoforms-Cox5a/Cox5b. COX subunit 5 (Cox5) is the homologue of subunit 4 (COX4) in mammals. In both organisms it exists in two isoforms, whose expression is regulated by oxygen. In yeast grown in normoxia, most COX holoenzyme contains the isoform Cox5a. In response to lowered oxygen availability, the isoform Cox5b is expressed. In addition, Cox5b/COX4-2 increases the efficiency of electrons transport through COX. By this mechanism conserved from lower to higher eukaryotes, cells are able to assemble, in response to hypoxia, a more efficient COX that will allow them to maximize ATP production and minimize ROS formation. Currently, the regulation of COX isozyme assembly is not completely disclosed. The unicellular yeast, S. cerevisiae is an excellent model for gaining insights into the functional basis of many human diseases particularly those stemming from impaired mitochondrial metabolism. To gain insights into the regulation of COX5b/COXIV-2, we used S. cerevisiae as a model system to explore new conserved regulatory mechanisms using either a genetic approach by isolating and investigating the respiratory deficiency suppressors of the normoxic cox5a deletion strain or a biochemical approach by studying the regulation of COX5b under different environmental stresses. We were able to discover novel regulatory mechanism for COX5b expression by reactive oxygen species, which enriches our understanding of the regulation of hypoxic MRC and could facilitate us to achieve the long term goal to develop therapeutic strategies to combat ischemic-reperfusion damage or mitochondrial deficiencies.

Keywords

mitochondria; cytochrome c oxidase; hypoxia; oxidative stress; reactive oxygen species; COX5; transcriptional regulation; allosteric regulation; aneuploidy

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