Leona Samson

Alkylating agents represent an abundant class of chemical DNA damaging agent in our environment and they are toxic, mutagenic, teratogenic and carcinogenic. Since we are continuously exposed to alkylating agents, and since certain alkylating agents are used for cancer chemotherapy, it is important to understand exactly how cells respond when exposed to these agents. The repair of DNA alkylation damage provides tremendous protection against the toxic effects of these agents and our aim is to understand the biology, the biochemistry, and the genetics of numerous DNA repair pathways that act upon DNA alkylation damage.

Organisms separated by enormous evolutionary distances employ similar proteins to protect against DNA damage, and we know that bacteria, yeast, and human cells induce the expression of specific sets of genes in response to such damage. Our studies on the response of Escherichia coli, Saccharomyces cerevisiae and human cells to alkylating agents have become intimately intertwined. Much of our previous work was based on the findings that bacterial DNA repair functions can operate in eukaryotic cells, and vice versa, i.e., eukaryotic DNA repair functions can operate in bacterial cells. We exploited this phenomenon to clone a large number of yeast, mouse and human DNA alkylation repair genes, and we are using these cloned genes to gain a thorough understanding of how eukaryotic cells respond to alkylating agents. Moreover, we have extended our alkylation toxicity studies from the cellular level to the whole animal level. Specifically, we have: (i) produced transgenic and knock-out mice with altered DNA repair capabilities and are now measuring their susceptibility to alkylation toxicity; and (ii) transferred DNA alkylation repair genes to bone marrow cells to determine whether such gene therapy could confer a useful level of extra resistance in the bone marrow of chemotherapy patients.

When cells are exposed to DNA damaging agents a signal is generated such that the transcription of various genes is altered. We have used Affymetrix oligonucleotide DNA chip analysis to monitor the transcriptional response of the entire S. cerevisiae genome, i.e., all 6,200 genes in response to a number of different alkylating agents. To our surprise, we have identified hundreds of responsive genes and have uncovered a hitherto unknown response that links ubiquitin-mediated protein degradation and DNA repair. We are currently exploring the biological roles that the large number of responsive genes plays in protecting cells against alkylation toxicity. Signals can also be generated, in cells explosed to alkylating agents, which trigger cell cycle checkpoint arrest or apoptosis. We are also dissecting the molecular mechanisms by which alkylating agents signal these very important downstream events.