Using live cell imaging to uncover how misrepair of double-strand breaks contributes to aging

2012 new Scholar Award in aging

Although aging is an immensely complicated process, one characteristic of aging is increased susceptibility to cancer, which can result from genetic instability. Our cells are exposed to DNA damaging agents daily, such as UV irradiation, and as we age the ability of our cells to repair these lesions becomes compromised. If DNA is not repaired accurately, then this can lead to genomic rearrangements and chromosomal abnormalities which are hallmarks of cancer cells. Underscoring the importance of DNA repair during aging, mutations in the RecQ family of related proteins are associated with three different cancer predisposition syndromes; Bloom, Werner, and Rothmund-Thomson syndromes. In addition to cancer observed in these patients, Werner and Rothmund-Thomson syndromes also cause premature aging (Bernstein et al. 2010).

We are studying how mutations in the RecQ family of proteins leads to these devastating diseases in the hopes that understanding the underlying mechanisms of these syndromes will elucidate how we age normally. To do this, we are using the model organism budding yeast, as a highly conserved and simple system where many different types of experiments can be readily accomplished. In fact, cells with mutations in the RecQ gene in yeast, SGS1, exhibit many characteristics observed in Bloom, Werner, or Rothmund Thompson syndromes, such as premature aging and genetic instability. The conservation between these proteins indicates how evolutionarily similar the DNA repair process is and suggests that what we learn in yeast is often directly applicable to human cells. We will then apply these finding to study aging in mammalian cells.

To understand how the RecQ genes contribute to aging, we are taking a novel imaging approach. To do this, we have fluorescently tagged the RecQ genes (i.e. with Yellow Fluorescent Protein) and can visualize their redistribution to DNA damage sites using fluorescent microscopy. In these same cells, we have an inducible DNA damage site that is also fluorescently tagged but in another color (i.e. Red Fluorescent Protein). We can now visualize the recruitment of the RecQ proteins to the DNA damage site or to multiple damage sites. Using this technique, we are asking how the RecQ proteins are recruited to DNA damage and how mis-regulation of their ability to go to DNA damage leads to genomic rearrangements. Our hypothesis is that the RecQ genes are involved in bringing broken DNA ends together for repair and we are specifically testing this model using the live-cell imaging approach described. Using what we discover, we will further understand how mutations in Sgs1 and its human orthologues (Bloom, Werner, and Rothmund-Thomson) contribute to DNA movement during DNA repair and how misregulation of this process can leads to premature aging and genomic rearrangements observed in tumor cells.