A proteomic atlas of senescence-associated secretomes for aging biomarker development Nathan Basisty, Abhijit Kale, Ok Hee Jeon, Chisaka Kuehnemann, Therese Payne, Chirag Rao, Anja Holtz, Samah Shah, Vagisha Sharma, Luigi Ferrucci, Judith Campisi, Birgit Schilling The Buck Institute for Research on Aging, Novato, California, United States of America; University of Washington, Seattle, Washington, United States of America; Intramural Research Program of the National Institute on Aging, NIH, Baltimore, Maryland, United States of America; Lawrence Berkeley Laboratory, University of California, Berkeley, California, United States of America This study presents the "SASP Atlas," a comprehensive proteomic database of soluble proteins and exosomal cargo SASP factors originating from multiple senescence inducers and cell types. The SASP is a complex secretory phenotype that includes cytokines, chemokines, growth factors, and proteases that initiate inflammation, wound healing, and growth responses in nearby cells. The SASP has been underestimated in complexity, and a small set of factors cannot explain the diverse phenotypes it produces in vivo. The study identifies several candidate biomarkers of cellular senescence that overlap with aging markers in human plasma, including GDF15, STC1, and SERPINs, which significantly correlated with age in plasma from a human cohort. The study also shows that the SASP is enriched for protein markers of human aging and proposes a panel of top SASP-based aging and senescence biomarker candidates. The study used a proteomic workflow to discover novel SASP factors. Senescent and quiescent primary human lung fibroblasts and renal cortical epithelial cells were induced to senesce by X-irradiation, inducible RAS overexpression, or atazanavir treatment. The secreted proteins and exosomes/EVs were separated by ultracentrifugation. The study identified between 441 and 1,693 secreted proteins per senescence inducer, a large fraction of which were significantly up- or down-regulated in the secretome following induction of senescence by IR, RAS, or ATV. The study also measured the secretion of known SASP factors in the fibroblast sSASP and eSASP, as well as the renal epithelial cell sSASP. These factors included CXCLs, HMGB1, IGFBPs, MMPs, LAMB1, and TIMPs. The study found that the sSASP is largely distinct in composition and regulation in fibroblasts and epithelial cells. The sSASP of irradiated fibroblasts and epithelial cells were largely distinct. Among the proteins increased in the sSASP of each cell type, 9%–23%A proteomic atlas of senescence-associated secretomes for aging biomarker development Nathan Basisty, Abhijit Kale, Ok Hee Jeon, Chisaka Kuehnemann, Therese Payne, Chirag Rao, Anja Holtz, Samah Shah, Vagisha Sharma, Luigi Ferrucci, Judith Campisi, Birgit Schilling The Buck Institute for Research on Aging, Novato, California, United States of America; University of Washington, Seattle, Washington, United States of America; Intramural Research Program of the National Institute on Aging, NIH, Baltimore, Maryland, United States of America; Lawrence Berkeley Laboratory, University of California, Berkeley, California, United States of America This study presents the "SASP Atlas," a comprehensive proteomic database of soluble proteins and exosomal cargo SASP factors originating from multiple senescence inducers and cell types. The SASP is a complex secretory phenotype that includes cytokines, chemokines, growth factors, and proteases that initiate inflammation, wound healing, and growth responses in nearby cells. The SASP has been underestimated in complexity, and a small set of factors cannot explain the diverse phenotypes it produces in vivo. The study identifies several candidate biomarkers of cellular senescence that overlap with aging markers in human plasma, including GDF15, STC1, and SERPINs, which significantly correlated with age in plasma from a human cohort. The study also shows that the SASP is enriched for protein markers of human aging and proposes a panel of top SASP-based aging and senescence biomarker candidates. The study used a proteomic workflow to discover novel SASP factors. Senescent and quiescent primary human lung fibroblasts and renal cortical epithelial cells were induced to senesce by X-irradiation, inducible RAS overexpression, or atazanavir treatment. The secreted proteins and exosomes/EVs were separated by ultracentrifugation. The study identified between 441 and 1,693 secreted proteins per senescence inducer, a large fraction of which were significantly up- or down-regulated in the secretome following induction of senescence by IR, RAS, or ATV. The study also measured the secretion of known SASP factors in the fibroblast sSASP and eSASP, as well as the renal epithelial cell sSASP. These factors included CXCLs, HMGB1, IGFBPs, MMPs, LAMB1, and TIMPs. The study found that the sSASP is largely distinct in composition and regulation in fibroblasts and epithelial cells. The sSASP of irradiated fibroblasts and epithelial cells were largely distinct. Among the proteins increased in the sSASP of each cell type, 9%–23%