Intellectual Merit: Chromosomes are duplicated during the S-phase of the cell cycle by the process of DNA replication. Once replication is initiated, two replication forks emerge from the origin of replication and progress in opposite directions along the DNA. When a moving replication fork encounters a DNA lesion on the template DNA strand, it can stall and subsequently collapse into a DNA double stranded break, which represents one of the most deleterious and mutagenic types of DNA damage for a cell. To stabilize, repair and restart replication at the damaged forks, a complex of DNA repair and replication proteins is recruited to stalled replication forks in order, thus protecting cells from genomic instability during the replication stage. Among these crucial proteins are a group referred to as the Fanconi anemia proteins (so-named because they were first discovered as being mutated in cases of Fanconi anemia), which play essential, yet poorly characterized, roles in maintaining replication fork integrity during every S-phase of the cell cycle. To explore how these proteins work, this research project will build on observations that DNA damage during S-phase triggers dissociation of a FANCD2-FANCI heterodimer into the monomeric subunits, which in turn are recruited separately to stalled replication forks where they fulfill different roles in fork repair and restart during S-phase. Research will test the question of whether FANCD2 and FANCI function separately to recruit distinct DNA repair protein complexes to sites of stalled replication forks as part of a process to restart replication. The results from this study will provide novel insights into the molecular mechanisms that ensure faithful and complete replication in the face of road-blocking DNA lesions.
Broader Impact: The proposed research is ideally suited for training future scientists, including two undergraduate research assistants, and two postdoctoral research associates. Lab scientists will acquire a broad set of experimental skills in the areas of molecular and cellular biology and biochemistry. Undergraduate assistants will be individually mentored and assigned integral roles in the research project. Encouraged to think creatively and independently, undergraduates will be involved in every step of developing important strategies and reagents as current and future research directions are being explored. To optimally prepare students and postdocs for future roles as scientists and lab leaders, several additional mechanisms are in place: All lab staff participate in, and present at, weekly joint lab meetings that bring together several other faculty and students. Undergraduate students and postdoctoral fellows are encouraged to present their research at national and international meetings in form of poster and oral presentations. Moreover, postdoctoral associates and undergraduate students have frequent opportunities to network with visiting scholars and are encouraged to initiate research collaborations to nurture their multidisciplinary research skills and experience. Results from this research project will be utilized in a new course, co-initiated by the PI on current and highly relevant topics of genomic stability for undergraduate and graduate students, that places strong emphasis on understanding experimental approaches, analyzing primary literature, designing future research goals and developing individual and group presentation skills. Lastly, scientific results of the project will be discussed and disseminated via publications and international meetings and contacts.
