The longstanding view has been that metabolism allows for cell division to take
place, but that metabolic processes do not actively promote cell division. I have recently
challenged this notion by identifying a unique gain-of-function metabolic mutant in the
budding yeast Saccharomyces cerevisiae. Moderate over-expression of Abf2p, a
conserved mitochondrial DNA (mtDNA) maintenance protein, increases the amount of
mtDNA by 100-150%. I have shown that cells moderately over-expressing Abf2p can
out-proliferate their wild type (WT) counterparts, initiate DNA replication sooner, and
increase in size faster than WT cells.
Yeast grown under certain conditions in continuous cultures become
synchronized with respect to their oxygen consumption, displaying distinctive oxidative
and reductive phases. In cells over-expressing Abf2p, the reductive phase is expanded
compared to that of WT cells. Since glutathione, the cell?s main redox buffer and sulfur
containing metabolite, peaks during this phase, I asked if sulfur metabolism was altered
in cells with more mtDNA.
Sulfur metabolite levels are increased ~40% in cells over-expressing Abf2p.
Furthermore, exogenous addition of various sulfur containing compounds, which is known to increase sulfur metabolic flux, caused WT cells to increase in size faster and
initiate DNA replication sooner, mimicking the phenotype seen in cells moderately overexpressing
Abf2p.
I then investigated possible interactions between sulfur metabolism enzymes and
the yeast Cdk, Cdc28p. Performing co-immunoprecipitation experiments, two enzymes
of the sulfur metabolic pathway were found to bind Cdc28p. One of these, Cys4p, lies at
the critical junction point between the pathways leading to the formation of glutathione
versus one carbon metabolism. The interaction of the enzymes with Cdc28p appears to
be dependent on progression through the cell cycle, and preliminary evidence suggests
that Cdc28p/Cys4p binding may peak at the G1/S transition of the cell cycle.
In summary, I have identified a unique gain-of-function metabolic mutant in S.
cerevisiae that leads to accelerated initiation of DNA replication. Sulfur metabolic flux
is up-regulated in cells over-expressing Abf2p, and exogenous sulfur sources added to
WT cultures phenocopied cells over-expressing Abf2p. Most importantly, I have shown
a physical interaction between sulfur metabolic enzymes and the Cdk driving the cell
cycle in yeast.
| Identifer | oai:union.ndltd.org:tamu.edu/oai:repository.tamu.edu:1969.1/ETD-TAMU-2009-12-7349 |
| Date | 2009 December 1900 |
| Creators | Blank, Heidi M. |
| Contributors | Polymenis, Michael |
| Source Sets | Texas A and M University |
| Language | English |
| Detected Language | English |
| Type | Book, Thesis, Electronic Dissertation, text |
| Format | application/pdf |
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