Hutchinson-Gilford Progeria Syndrome (HGPS) is a rare, fatal autosomal dominant genetic condition characterized by many characteristics of accelerated aging in children. Children with HGPS die at an average age of fourteen, usually from heart disease. It has been known for some time that the syndrome is most commonly caused by a specific point mutation in the LMNA gene which normally codes for lamin A and its splice variant lamin C. The LMNA mutation commonly associated with HGPS leads to increased usage of a cryptic splice site which leads to the production of a truncated, farnesylated form of lamin A referred to as progerin. Quite interestingly, progerin is expressed at very low levels in healthy individuals and appears to play a role in the normal aging process. Lamin A normally serves as an important component of the nuclear lamina, a structure that provides mechanical support for the nucleus and regulates several critical cellular processes including a number of DNA repair pathways. In HGPS, the impact of overexpression of progerin on nuclear architecture and genomic stability is profound. One reported consequence of progerin expression is an accumulation of DNA double- strand breaks (DSBs). To our knowledge, however, the effect that expression of progerin or other mutated forms of lamin A has on the precise manner in which a DSB is repaired in the human genome has never been investigated in detail. This gap in our knowledge is significant since corruption of DNA repair is commonly viewed as playing a key role in the aging process. Over the past two decades, we have developed and productively used a model system for studying DSB repair, homologous recombination, and related DNA transactions in the chromosomes of human cells. We have engineered DNA constructs containing a mutated selectable marker gene and a closely linked sequence that can restore function to the marker through recombination. Constructs are stably transfected into cell lines of interest. A DSB can be introduced into the marker gene by transient expression of endonuclease I-SceI and so we can recover a variety of DSB repair events that restore marker function by applying genetic selection. We will apply our system to compare and contrast DSB repair events in normal cells versus cells that overexpress progerin or express a related variant form of lamin A. Our work will be the first in-depth investigation into how expression of progerin impacts DSB repair pathway choice as well as the nature of individual repair events at the nucleotide level. We will focus on the questions of whether expression of progerin reduces the stringency of recombinational repair, enabling genetic exchange between imperfectly matched sequences, and whether progerin inhibits precise joining of DNA ends. Either of these potential impacts of progerin expression would likely destabilize the genome. Our studies will provide novel information about the aging process, information that will enhance our understanding of progeria and normal aging.
A protein named ?progerin? has been implicated in the normal aging process and, when expressed at high levels, progerin causes a disease associated with premature aging. We will explore the possibility that progerin and related proteins have negative impacts on a cell?s ability to repair damage to chromosomes. By enhancing our understanding of how progerin and related proteins may interfere with the maintenance of chromosome integrity, we will gain insight into mechanisms of normal and abnormal human aging and, possibly, how to ameliorate the impacts of aging.
