The remarkable increase in human longevity over the last century has, unfortunately, increased the risk of developing a number of age-related conditions, which underscores the need to develop strategies to extend human healthspan. Cells recycle their damaged organelles and protein aggregates via autophagy whereby cytoplasmic components are sequestered within autophagosomes (APh) and targeted for lysosomal degradation. Briefly, class III PI3K, ULK1 kinase, and a conjugation cascade involving core autophagy-related (ATG) proteins, ATG7, ATG5, ATG12, ATG16L1 and ATG8/light chain 3 (LC3) regulate autophagosome (APh) formation by lipidating cytosolic LC3-I into APh-bound LC3-II. Aging associates with autophagy failure, which is a critical factor in the development of age-associated conditions. How autophagy decreases with age remains unclear. While sustained mTOR activation, altered membrane lipid composition, and acetylation of core ATG proteins each contribute to autophagy failure; none of these mechanisms can completely explain why ATG proteins decrease with age. We have found that LC3, a structural component of APh, binds to Atg16l1 mRNA. LC3 has recently been shown to bind and enhance fibronectin mRNA translation ? thus revealing a novel role of LC3 in RNA metabolism. Our preliminary analyses revealed that starvation, which activates autophagy, leads to increased LC3-Atg16l1 mRNA binding. Livers knocked out for Atg7 (that do not form LC3-II), and livers from aged mice, each display loss of LC3-Atg16l1 mRNA binding, suggesting that LC3-II, and not LC3- I, binds to Atg16l1 mRNA. In addition, overexpression of LC3 enhances ATG16L1 protein half-life. Furthermore, Atg16l1 mRNA sequence analysis revealed presence of several AU-rich elements (ARE), which are cis-acting sequences known to facilitate binding of RNA to specific proteins. These observations have led to our hypothesis that LC3 binds to Atg16l1 mRNA in a feed-forward regulatory loop that stabilizes Atg16l1 mRNA, and thereby maintains ATG16L1 protein levels and autophagy. We hypothesize further that disruption of this regulatory loop contributes to autophagy failure. The overall goal of this proposal is to: 1) examine whether age-associated decrease in LC3-Atg16l1 mRNA binding leads to decreased Atg16l1 mRNA levels and autophagy failure, 2) determine the mechanism of LC3- mediated Atg16l1 mRNA stability by using novel LC3 mutants in vitro, and a novel RRRLC3QQQ mutant mouse wherein a triple arginine (RRR) motif in LC3, which we propose allows LC3-Atg16l1 mRNA binding, has been inactivated, and 3) test whether genetic reconstitution of LC3-II in aged livers can restore Atg16l1 mRNA levels, autophagy, and lipohomeostasis. Our studies will provide new mechanistic insight into how autophagy decreases with age, and provide a novel strategy to prevent the decline of this important quality control pathway with age. Significance: Aged individuals are at increased risk for developing a number of diseases, neurodegeneration, metabolic syndrome, and cancers, to mention a few. Studies of the loss of autophagy in young mice or genetic overexpression of specific Atg genes in mice have shown the critical role of autophagy in disease prevention. How autophagy decreases with age remains unclear. Our studies will reveal a new mechanism of regulating autophagy by enhancing the stability of an Atg mRNA, Atg16l1, by an ATG protein, LC3. Understanding how autophagy decreases with age is critical for the development of new strategies to extend human healthspan ? the number of years one remains healthy and active.
Aged individuals are at increased risk for developing a number of diseases, neurodegeneration, metabolic syndrome, and cancers, to mention a few. Age-associated loss of a cellular quality control pathway, autophagy, plays a significant role in development of these diseases. How autophagy decreases with age remains unclear. The studies enlisted in this application will reveal a new mechanism of regulating autophagy through stabilization of autophagy gene transcripts, which will throw light on why autophagy decreases with age. Our studies will be critical for the development of new strategies to extend human healthspan through restoration of autophagy.
| Toledo, Miriam; Batista-Gonzalez, Ana; Merheb, Emilio et al. (2018) Autophagy Regulates the Liver Clock and Glucose Metabolism by Degrading CRY1. Cell Metab 28:268-281.e4 |
| Batista-Gonzalez, Ana; Singh, Rajat (2017) Lysosomal function in ?-cell survival during glucolipotoxicity. Ann Transl Med 5:471 |
