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Probing Iron Accumulation in Sacchromyces cerevisiae Using Integrative Biophysical and Biochemical Techniques

Iron is an essential element for life. It is involved in a number of biological
processes, including iron sulfur (Fe/S) cluster assembly and heme biosynthesis. However
it is also potentially toxic due to its ability to induce formation of reactive oxygen
species (ROS) via Fenton chemistry. Therefore its uptake, trafficking and utilization
must be regulated to avoid its toxicological effect. It has been recently discovered that
Fe/S cluster biosynthesis machinery plays a key role in the cellular iron regulation and
its disruption leads to impaired iron regulation and iron accumulation within
mitochondria.
The iron accumulation resulted from impaired Fe/S cluster assembly in the
eukaryotic model organism Saccharomyces cerevisiae (baker’s yeast) was studied.
Various biophysical (e.g. Mössbauer, EPR, UV-vis spectroscopy) and biochemical (e.g.
Western blots, PCR, enzyme activity assay, etc.) techniques were used to characterize the
iron content in yeast mitochondria isolated from several mutants strains. In these mutants
one of the proteins involved in Fe/S cluster biosynthesis (Yah1p and Atm1p) is mutated and iron regulation and metabolism are disrupted. By integrating the results obtained
from these different methods, it was determined that excess iron accumulates in the
mutant mitochondria as inorganic phosphate Fe(III) nano-particles exhibiting
superparamagnetic behaviors. Oxygen is required for iron accumulation and nanoparticle
formation. The Fe(III) nano-particles can be chemically reduced to Fe(II) then largely
exported from the mitochondria.
These biophysical and biochemical methods were also used to examine the iron
distribution in whole yeast cells of the Aft1-1up strain in which iron regulon genes are
constitutively activated and compared to that of Yah1p-depleted and wild type yeast.
Constitutive activation of iron regulon genes does not alter the cellular iron distribution
significantly. However disruption of Fe/S cluster assembly by Yah1p depletion causes
dramatic cellular iron redistribution: the vacuolar iron is largely evacuated and most of
the cellular iron probably precipitates in mitochondria as Fe(III) nanoparticles. The
results provide novel insights into iron trafficking and possible signal communications
between organelles within cells.

Identiferoai:union.ndltd.org:tamu.edu/oai:repository.tamu.edu:1969.1/ETD-TAMU-2010-12-8743
Date2010 December 1900
CreatorsMiao, Ren
ContributorsLindahl, Paul A.
Source SetsTexas A and M University
Languageen_US
Detected LanguageEnglish
Typethesis, text
Formatapplication/pdf

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