Research Projects

Protein misfolding and aggregation contribute to major age-related diseases such as neurodegeneration, diabetes, cardiovascular disease, and cancer conditions that increasingly burden India’s rapidly aging population. With India’s elderly expected to exceed 300 million by 2050, these diseases pose critical challenges to healthcare and economic systems. Aging disrupts proteostasis, leading to the accumulation of damaged or misfolded proteins. This disruption contributes significantly to functional decline and the development of age-related diseases; thus, loss of proteostasis is considered one of the hallmarks of aging. This suggests that boosting proteostasis is one of the ways to extend healthy lifespan. Cells maintain protein quality control (PQC) systems to monitor and protect the proteome to maitain proteostasis. In PQC, molecular chaperones assist in protein folding and prevent aggregation, whereas degradation systems eliminate misfolded proteins. However, PQC tends to decline with aging, leading to an increase in protein misfolding and aggregation, which is a common feature of many age-related diseases. Protein disaggregases, which reverse protein aggregation, are the poorly characterized components of the PQC in animal systems. To identify the molecular machinery responsible for the animal disaggregase activity, we developed a C.elegans model that expresses a proteostasis sensor (modified firefly luciferase fused to green fluorescent protein, DMFluc:GFP) in muscle and neuronal tissues. This sensor forms aggregates upon heat stress and resolubilizes with recovery. However, with age, these heat-induced aggregates fail to resolubilize completely. This sensor also reports tissue-specific differences in the proteostasis capacity. Intriguingly, it seems these heat-induced aggregates are not degraded by the protein degradation systems (preliminary observations), suggesting the existence of active disaggregation machinery. The insulin-like pathway regulates activities of HSF-1 and DAF-16 (FOXO) transcription factors that promote lifespan and PQC in diverse systems. Surprisingly, we found that the heat-induced DMFluc aggregation is reversible only in daf-16 but not in hsf-1 mutant animals. While heat-induced protein aggregation is effectively resolved in young wild-type and daf-16 mutant animals, this recovery is compromised in aged animals. In contrast, hsf-1 mutants are unable to recover from aggregation at any age. This suggests that the proteostasis environment in young hsf-1 mutants resembles that of aged wildtype and daf-16 mutants, particularly regarding protein disaggregation capacity. This indicates that HSF-1 and DAF-16 function differently in managing protein aggregation. In fact, it has already been reported that HSF-1 promotes disaggregation (primary pathway), whereas DAF-16 promotes aggregation (secondary pathway) as part of cellular stress response. However, molecular identity of the disaggregase components is unknown. This proposal is aimed at identifying such modifiers of protein aggregates by mass-spectrometry (MS)-based proteomics. The DMFluc:GFP will be isolated from young and aged wildtype, hsf-1, and daf-16 mutant animals before and after heat stress and recovery by coimmunoprecipitation. The interactors will be identified by MS. This approach will overcome the efforts to identify novel modifiers of protein aggregation, including disaggregases. In addition, total protein abundance changes will also be identified before and after heat stress in young and aged animals. Comparison of this abundance data with the available transcriptomic data from hsf-1 and daf-16 mutant animals will help in identifying unique as well as shared targets of HSF-1 and DAF-16. Furthermore, these potential candidates will be validated using model disease aggregation proteins. We assume these proteins also modulate PQC in a better way to promote and prolong a healthy lifespan.