Faculty Mentor
Daniel Ginsburg
Major/Area of Research
Biomedical Sciences
Description
Chromatin is the complex of proteins and DNA that makes up eukaryotic
chromosomes. Chromatin helps the DNA fit inside the nucleus as well as
regulating what can access the DNA. Replication, repair, and transcription
are all impeded by chromatin. These processes only occur efficiently
when chromatin is disassembled. Just as chromatin limits how proteins
can access the DNA, we hypothesized that it would also protect DNA. We
used the yeast Saccharomyces cerevisiae to study how chromatin affects
cellular sensitivity to the DNA alkylating agent methyl methanesulfonate
(MMS). MMS alkylation can lead to DNA mismatches, nicks, and double
strand breaks, resulting in mutation. Using growth assays, we found that
active transcription significantly increases the sensitivity of a gene to MMS.
Consistent with this, we found that increased chromatin compaction due to
nocodazole-mediated metaphase arrest reduced the effects of MMS. Because
transcription led to an increase in MMS-mediated DNA damage, we proposed
that transcribed genes would be subject to increased damage under
normal growth conditions, requiring recruitment of DNA repair factors. We
found by chromatin immunoprecipitation that the Rad51, Rad53, and Rad18
DNA repair proteins are recruited to the GAL1 gene during transcription,
although their occupancy was not stimulated by MMS treatment. This suggests
that there is enough damage occurring during normal transcription
to require DNA repair factors. Together, these results support our model that
chromatin helps protect DNA from damage in addition to regulating other
DNA-based processes.
Included in
Chromatin Condensation Protects DNA from MMS Damage
Chromatin is the complex of proteins and DNA that makes up eukaryotic
chromosomes. Chromatin helps the DNA fit inside the nucleus as well as
regulating what can access the DNA. Replication, repair, and transcription
are all impeded by chromatin. These processes only occur efficiently
when chromatin is disassembled. Just as chromatin limits how proteins
can access the DNA, we hypothesized that it would also protect DNA. We
used the yeast Saccharomyces cerevisiae to study how chromatin affects
cellular sensitivity to the DNA alkylating agent methyl methanesulfonate
(MMS). MMS alkylation can lead to DNA mismatches, nicks, and double
strand breaks, resulting in mutation. Using growth assays, we found that
active transcription significantly increases the sensitivity of a gene to MMS.
Consistent with this, we found that increased chromatin compaction due to
nocodazole-mediated metaphase arrest reduced the effects of MMS. Because
transcription led to an increase in MMS-mediated DNA damage, we proposed
that transcribed genes would be subject to increased damage under
normal growth conditions, requiring recruitment of DNA repair factors. We
found by chromatin immunoprecipitation that the Rad51, Rad53, and Rad18
DNA repair proteins are recruited to the GAL1 gene during transcription,
although their occupancy was not stimulated by MMS treatment. This suggests
that there is enough damage occurring during normal transcription
to require DNA repair factors. Together, these results support our model that
chromatin helps protect DNA from damage in addition to regulating other
DNA-based processes.