PROJECT SUMMARY/ABSTRACT Our overarching goals are: i. to define the mechanism(s) of ATM activation by the Mre11 complex; ii. to define the role(s) of the Mre11 complex in DNA replication; iii. To define the role(s) of the Mre11 complex in response to DNA replication stress. As part of the emphasis of DNA replication stress, our focus includes RTEL1 which acts to mitigate replication stress at the telomere. A combination of yeast and mouse genetics, biochemistry, and structural biology are employed to address these issues. Our previous work on these topics led to substantial scientific progress and insight regarding processes relevant to human health. Specific areas of inquiry are: ? Genetic screens in yeast and mining of cancer genomic data revealed separation of function Rad50 mutations that affect ATM dependent DNA damage signaling without affecting the DNA repair functions of the Mre11 complex. These mutations constitute a resource for deciphering the mechanism by which the Mre11 complex activates the ATM kinase to initiate DNA damage signaling. ? We have modeled three recurrent tumor borne mutations in mice that were confirmed to be hypomorphic for ATM activation but proficient in DNA repair. These animal models offer a unique opportunity to delineate Mre11 complex-dependent ATM functions. ? We have identified factors that function at the DNA replication fork in a manner that depends on the Mre11 complex. An important focus of our laboratory is to understand the functional role(s) of those factors. Defects in factors that promote accurate DNA replication are highly correlated with human disease, and so understanding this fundamental process is an important priority in our work. ? Whereas the Mre11 complex functions at the replication fork, RTEL1 is a helicase that promotes accurate replication of telomeric DNA. We have discovered that RTEL1 influences the abundance and disposition of a long non coding RNA, called TERRA, that is transcribed from the subtelomeric regions of all eukaryotes. Our goal in this aspect of our work is two fold. First, to understand the role of RTEL1 in maintaining telomere stability. Second, to use that information to shed light on the function of TERRA, which is a long standing question.