Date of Award

Spring 4-16-2014

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biology

First Advisor

John E. Houghton

Second Advisor

Susanna F. Greer

Third Advisor

Irene T. Weber

Abstract

Chronic exposure of heavy metals is highly correlated with the epidemic of degenerative disease, nephrotoxicity, cancers and aging, while the acute response of cells to the same heavy metals provides some nuanced insights into how cells are able to handle environmental insults, and perhaps characterize specific triggers of the process, itself. Some heavy metals, such as copper, trigger an immediate accumulation of reactive oxygen species (ROS), which impair vital cellular functions by oxidative stresses; which can lead to the onset of programmed cell death, or apoptosis, which becomes an inevitable fate once the damage is too disseminative to be recovered. Other heavy metals, such as cadmium, appear to trigger similar apoptotic responses in the cell –even before ROS increases to unmanageable levels. In either instance, however, before undergoing apoptosis, there are two cellular defensive mechanisms that are able to eliminate the metal-induced oxidative stresses: 1) the neutralization of anti-oxidants, and 2) the removal of the harmful substances through a series of self-cleaning mechanisms. We have used Saccharomyces cerevisiae, or baker’s yeast, as a model organism to demonstrate the response of cells to the presence of heavy metals. In so doing, we highlight pertinent aspects of the metabolic transcriptome response of these unicellular organisms to the presence of these metals, such as changes in expression of genes involved in the pentose phosphate pathway (PPP), which facilitates the reduction of oxidative glutathione, or induction of the genes most commonly associated with autophagy. These findings serve to indicate the protective mechanisms that are triggered upon metal exposures in yeast. Curiously, we also discovered that the autophagic response may be duplicitous, in that while the autophagic process can be cyto-protective it can also enhance the self-destructive mechanisms of the apoptotic response, indeed it is appears to be a requisite part of that response. Whether or not the cells respond to the cellular stress by autophagy or apoptosis appears to be “decided” by whether or not a full-blown autophagic response to cellular stressors (which can be independently induced by the drug, rapamycin) is initiated before the same autophagic process is able to trigger activation of a caspase-induced apoptosis. In addition, in order to monitor the autophagic process more carefully, we have developed a cytometric methodology to assess the autophagy flow, that is less labor-intensive and more dynamic than the traditional Western blot-based method. In so doing we have been able to decipher the factors of cell fate decision with heavy metal-induced oxidative stress in S. cerevisiae.

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