Piening, Brian (2013) Novel connections between DNA replication, telomere homeostasis and the DNA damage response revealed by a genome-wide screen for TEL1/ATM interactions in Saccharomyces cerevisiae. PhD thesis, Fred Hutchinson Cancer Research Center / University of Washington.
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Abstract
Tel1p is the budding yeast ortholog of the mammalian tumor suppressor and DNA damage response (DDR) kinase ATM. However, tel1-deletion cells, unlike ATM-deficient cells, do not exhibit sensitivity to DNA damaging agents, but do display shortened (but stably maintained) telomere lengths. Neither the extent to which ATM/Tel1p functions in the DDR nor the mechanism by which ATM/Tel1p contributes to telomere metabolism is well-understood. In this dissertation, I present our large-scale transcriptional profiling of normal and ATM-deficient lymphoblast cell lines in response to ionizing radiation (IR). From these results, we make the surprising observation that ATM-deficient cells exhibit no significant defects in IR-induced gene expression, which along with work from others suggests that significant redundancy exists in the DNA damage response, and may be an explanation for the relative DNA damage insensitivity in tel1-deletion yeast cells. To address this question, I performed a comprehensive genome-wide screen for genetic interactions with tel1-deletion that cause sensitivity to MMS and/or ionizing radiation, along with follow-up characterizations of the 13 interactions yielded by this screen. Surprisingly, many of the tel1-deletion interactions that confer DNA damage sensitivity also exacerbate the short telomere phenotype, suggesting a connection between these two phenomena. Restoration of normal telomere length in the tel1-deletion xxx-deletion mutants results in only minor suppression of the DNA damage sensitivity, demonstrating that the sensitivity of these mutants must also involve mechanisms independent of telomere length. In support of a model for increase replication stress in the tel1-delta xxx-delta mutants, I show that depletion of dNTP pools through pre-treatment with hydroxyurea renders tel1-delta cells (but not wild-type) MMS-sensitive, demonstrating that under certain conditions, Tel1p does indeed play a critical role in the DDR.ATM-deficient cells exhibit no significant defects in IR-induced gene expression, which along with work from others suggests that significant redundancy exists in the DNA damange response, and may be an explanation for the relative DNA damage insensitivity in tel1-delta yeast cells. To address this question, I performed a comprehensive genome-wide screen for genetic interactions with tel1-delta that cause sensitivity to MMS and/or ionizing radiation, along with follow-up characterizations of the 13 interactions yielded by this screen. Surprisingly, many of the tel1-delta interactions that confer DNA damange sensitivity also exacerbate the short telomere phenotype, suggesting a connection between these two phenomena. Restoration of normal telomere length in the tel1-delta xxx-delta mutants results in only minor suppression of the DNA damage sensitivity, demonstrating that the sensitivity of these mutants must also involve mechnaisms independent of telomere length. In support of a model for increase replication stress in the tel1-delta xxx-delta mutants, I show that depletion of dNTP pools through pre-treatment with hydroxyurea renders tel1-delta cells (but not wild-type) MMS-sensitive, demonstrating that under certain conditions, Tel1p does indeed play a critical role in the DDR.
| Item Type: | Thesis (PhD) |
|---|---|
| Subjects: | Molecules > Chromosomes Cellular and Organismal Processes > Genetic processes > DNA damage and repair Cellular and Organismal Processes > Genetic processes > DNA replication |
| Depositing User: | Craig Johansen |
| Date Deposited: | 24 Sep 2014 21:30 |
| Last Modified: | 24 Sep 2014 21:30 |
| URI: | http://authors.fhcrc.org/id/eprint/618 |
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