Shedding light on the link between the actin cytoskeleton and stress response in Saccharomyces cerevisiae : a thesis submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy in Biochemistry, Massey University, Auckland, New Zealand
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Date
2012
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Massey University
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Abstract
Understanding how eukaryotic cells adapt to stress remains a fundamental
question in biology. One important stress which affects all cells is
amino acid (AA) starvation. Upon perception of AA starvation the Gcn2
protein binds an effector protein complex consisting of Gcn1 and Gcn20.
Subsequently, Gcn2 becomes activated and phosphorylates the eukaryotic
translation initiation factor 2. This leads to a reduced global protein synthesis
and simultaneously to an increased translation of Gcn4, a transcriptional
activator of genes necessary for overcoming stress. This signaling pathway
is called general amino acid control (GAAC) in yeast. Failing to activate this
signaling cascade impairs the starvation response and cellular growth.
Yih1 inhibits Gcn2 by competing with Gcn2 for Gcn1 binding, consequently
leading to an impaired stress response. However, Yih1 is not a
general inhibitor of Gcn2 but only impedes Gcn2 activation upon release
from the cytoskeleton protein actin. Our understanding of the role of actin
in Gcn2 signaling and Yih1 itself is limited. Also, the circumstances under
which Yih1 is released from actin are unknown. Thus, the scope of this study
is to elucidate the link between the actin cytoskeleton and stress response.
To achieve this goal, actin mutants were screened for an impaired ability to
overcome starvation. Out of 24 mutant strains five exhibited an impaired
stress response as indicated by sensitivity to AA analogues, which could
be reverted upon Gcn4 induction, and sensitivity to AA imbalance. One of
this actin mutations has been proposed to affect the GAAC via an impaired
Yih1-actin binding. Another mutation appears to weaken the actin-eEF1A
interaction therefore promoting eEF1A mediated Gcn2 inhibition. For the
remaining 3 actin mutations the mechanism might be at the transcriptional
or translational level. These findings show that actin mutations do affect the
GAAC at multiple levels. In addition, the cyclin dependent kinase Cdc28
has been identified as a novel interaction partner of Yih1 and it has been
speculated that a link between Yih1 and bud emergence might exist.
The results from this study achieved to shed light on the link between
Yih1, the GAAC and the actin cytoskeleton. Yih1 has been placed in the
midst of highly regulated and diverse cellular processes, which emphasizes
the interconnection that exist between cellular pathways and the likely importance
of Yih1 in the cell.
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Keywords
Saccharomyces cerevisiae, Actin cytoskeleton, Stress response, Stress physiology, Amino acid starvation, General amino acid control