PROJECT SUMMARY
Cellular senescence is a stable form of cell cycle arrest associated with inflammatory responses. Senescent
cells accumulate in aged and diseased tissues and are considered as one of the major sources contributing to
chronic inflammation that is implicated in most, if not all, age-associated disorders. Consistent with this notion,
genetic or pharmaceutical clearance of senescent cells extend lifespan and healthspan of mice.
Senescent cells secret a large array of pro-inflammatory cytokines, chemokines, growth factors, and
proteases, collectively referred to as senescence-associated secretory phenotype (SASP). The SASP program
alters tissue microenvironment and recruits immune cells, ultimately leading to inflammation. Our group recently
showed that senescent cells exhibit genomic DNA in the cytosol, which is interpreted by the cells as a “danger
signal” by triggering the innate immunity cytosolic DNA sensing cGAS-STING pathway that promotes the SASP
program of senescence. These findings have been independently reproduced by several laboratories, and
collectively the cGAS-STING pathway is considered as a central mechanism for the SASP program.
A major unaddressed question in senescence is the genetic origin of cytosolic genomic DNA. Our imaging
results suggest that the cytosolic DNA is derived from fragments of chromatin, mediated by nucleus-to-cytoplasm
trafficking, via nuclear membrane blebs that partition into the cytoplasm. But which parts of the genome are
shuttled to the cytoplasm? What is the chromatin status of those regions? Does the genome lose genes? These
questions require unbiased sequencing approaches to address. This application proposes two novel strategies
to sequence cytoplasmic DNA in senescent cells. First, we aim to identify cGAS-associated cytosolic DNA in
senescence, by performing cGAS DNA-immunoprecipitation. Second, we aim to biochemically fractionate the
DNA from the cytoplasm of senescent cells. The DNA samples will be subjected to next-gen sequencing and
computational analyses to explore the chromatin marks and gene expression status. These results will permit
us to directly manipulate the genomic DNA to inquire the functional consequences of cells undergoing genomic
DNA trafficking to the cytoplasm.
This study will help the senescence field understand a critical mechanism underlying senescence-
associated inflammation, and may reveal previously unknown knowledge of the genetic alterations of
senescence and aging. This study has the potential to facilitate new approaches to target and inhibit chronic
inflammation to promote healthy aging and to suppress age-associated diseases.