In StemProteostasis, we found that proteasome inhibition impairs diverse processes required for hESC identity (Saez et al, Scientific Reports, 2018). To identify endogenous targets of the proteasome, we focused on E2 ubiquitin-conjugating enzymes and E3 ubiquitin ligases, which mark specific targets for proteasomal degradation. By integrating proteomics and transcriptomics data, we defined up-regulated E3s in hESCs that are required for pluripotency. Among them, we found that UBR5 suppresses proteostasis collapse in stem cells and can be applied into HD models to prevent disease-related changes (Koyuncu et al, Nature Communications 2018). In addition, we discovered that the E2 enzyme UBE2K promotes proteasomal degradation of histone H3 and its trimethylase SETDB1 in hESCs, maintaining low levels of H3K9me3 and neurogenic potential (Fatima et al, Communications Biology, 2020). To define how the proteasome regulates organismal aging, we performed a comprehensive analysis of ubiquitination changes through the entire proteome of C. elegans during aging. By integrating data from animals with a defective proteasome, we identified proteasomal targets that accumulate with age due to decreased ubiquitination and subsequent degradation. Notably, lowering their protein levels prolonged longevity, whereas preventing their proteasomal degradation shortened lifespan. Among them, we found the intermediate filament, IFB-2, and EPS-8, a modulator of RAC signaling. While increased IFB-2 levels promoted loss of intestinal integrity and bacterial colonization, upregulated EPS-8 hyperactivated RAC in muscle and neurons leading to alterations in actin cytoskeleton. Thus, we demonstrated that age-related changes in targeted degradation of structural and regulatory proteins across tissues determine longevity (Koyuncu et al, Nature 2021).
Beyond the proteasome, we defined other upregulated proteostasis mechanisms in hESCs/iPSCs and proved their role as anti-aging factors. First, we discovered that hESCs have upregulated activity of the TRiC/CCT complex, a chaperonin that facilitates the folding of 15% of the human proteome. We observed that somatic overexpression of CCT8, a TRiC/CCT subunit, extends C. elegans lifespan and ameliorates HD-related changes (Noormohammadi, Nature Communications, 2016). In hESCs, we also found increased expression of specific RNA-binding proteins (i.e. cold-shock enzymes) required for stem cell function (Lee et al, Nature Communications, 2017). Notably, these RNA-binding proteins determined C. elegans longevity through their activity in germline stem cells (Lee et al, Nature Metabolism 2019). Importantly, we found that a collapse in germline proteostasis leads to systemic mitochondrial alterations in tissues such as muscle and neurons through long-range signals. Subsequently, these alterations increased aggregation of HD and ALS-related proteins in somatic tissues such as neurons (Calculli & Lee et al, Science Advances, 2021).
Finally, we examined whether proteostasis regulates viability of plant stem cells in Arabidopsis thaliana. In contrast to their differentiated counterparts, root stem cells could prevent the accumulation of aggregated proteins even under proteotoxic stress conditions such as heat stress or proteasome inhibition. Notably, endogenous high levels of the TRiC/CCT complex were required for plant stem cell maintenance and their remarkable ability to suppress protein aggregation. Moreover, overexpression of CCT8 was sufficient to ameliorate protein aggregation in differentiated cells and confer resistance to proteotoxic stress in plants. Together, our results established enhanced proteostasis in stem cells as an important requirement for plants to persist under extreme environmental conditions and reach extreme long ages (Llamas et al, Aging Cell 2021).
