Subject Area

Biochemistry and Molecular Biology

Description

Mitochondria are essential organelles in both yeast and human cells due to their role in metabolism, ATP production via the oxidative phosphorylation (OXPHOS) pathway, and other regulatory cellular processes. Using yeast as a model organism to study mitochondrial function in a chronological lifespan assay allows experiments to be conducted over a shorter timeframe and allows for connections to be made to human cells. Our aim is to investigate how exogenous copper in the mitochondria of yeast affects the production of reactive oxygen species (ROS), protein expression, and enzyme activity during yeast lifespan. Small amounts excess copper added to growth media (0.25 mM copper sulfate in restricted nutrient media) extend yeast chronological lifespan, but yeast lifespan is reduced when added copper levels are increased to 2.0 mM copper sulfate or higher. These results indicate that low levels of exogenous copper in the media is beneficial for yeast in these restricted media conditions. To extend these findings, we assessed how added copper changes mitochondria within these cells over the course of the yeast lifespan (14 days growth). Using MitoTracker Green and fluorescence detection we showed an increase in mitochondria in copper treated cells. This is consistent with previous studies showing mitochondria in mammal and yeast cells contain a labile copper pool located in the matrix, which is used in the metalation of the copper containing enzyme of the OXPHOS pathway Complex IV, Cytochrome C Oxidase (CcO), and superoxide dismutase (Sod1p). Our most recent work focuses on assessing CcO protein complex expression during yeast lifespan, specifically looking at cytochrome c oxidase subunits using western blotting; and assessing Sod1p activity using in-gel activity assays. The results of this research allow us to better understand the role of copper in mitochondrial activity across the lifespan of yeast.

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The effect of low concentrations of copper on mitochondria and activity in yeast cells

Mitochondria are essential organelles in both yeast and human cells due to their role in metabolism, ATP production via the oxidative phosphorylation (OXPHOS) pathway, and other regulatory cellular processes. Using yeast as a model organism to study mitochondrial function in a chronological lifespan assay allows experiments to be conducted over a shorter timeframe and allows for connections to be made to human cells. Our aim is to investigate how exogenous copper in the mitochondria of yeast affects the production of reactive oxygen species (ROS), protein expression, and enzyme activity during yeast lifespan. Small amounts excess copper added to growth media (0.25 mM copper sulfate in restricted nutrient media) extend yeast chronological lifespan, but yeast lifespan is reduced when added copper levels are increased to 2.0 mM copper sulfate or higher. These results indicate that low levels of exogenous copper in the media is beneficial for yeast in these restricted media conditions. To extend these findings, we assessed how added copper changes mitochondria within these cells over the course of the yeast lifespan (14 days growth). Using MitoTracker Green and fluorescence detection we showed an increase in mitochondria in copper treated cells. This is consistent with previous studies showing mitochondria in mammal and yeast cells contain a labile copper pool located in the matrix, which is used in the metalation of the copper containing enzyme of the OXPHOS pathway Complex IV, Cytochrome C Oxidase (CcO), and superoxide dismutase (Sod1p). Our most recent work focuses on assessing CcO protein complex expression during yeast lifespan, specifically looking at cytochrome c oxidase subunits using western blotting; and assessing Sod1p activity using in-gel activity assays. The results of this research allow us to better understand the role of copper in mitochondrial activity across the lifespan of yeast.