Investigating the role of the CCT chaperonin in stem cell identity and aging

With the constant increase in the age of the population in many countries of the world, understanding the molecular mechanisms of aging is of central importance. This increase in the age of societies in developed countries has been achieved through improvements in healthcare and drug development. However, aging is associated with a myriad of diseases that not only affect the quality of life for the elderly, but also impose a major challenge on the socio-economic and healthcare systems. Thus, gaining a better understanding of the aging process at the molecular level could lead to novel therapies against aging and age-associated diseases. Functional decay of somatic stem cells, which leads to hampered tissue regeneration is one of the main hallmarks of aging. Human embryonic stem cells (hESCs) can replicate indefinitely while maintaining their undifferentiated state and, therefore, are immortal in culture. This capacity is among others dependent on enhanced protein homeostasis (proteostasis) mechanisms, since any imbalance in protein quality control could potentially compromise the resulting lineage of cells. Decline in the proteostasis is also known as a main contributor to aging. Thus, studying proteostasis of hESCs is a novel paradigm, that will not only help us gain a better understanding of hESC biology, but could also provide a link between hESC immortality, organismal longevity and stress resistance. In the frame of this study, we aimed to investigate the regulation of proteostasis in hESCs and its relevance to stemness, stress resistance, aging and age-associated diseases. We tried to define key candidates responsible for enhanced proteostasis in hESCs by comparing hESCs and differentiated cells using quantitative proteomics and RNA-sequencing, and were able to identify several proteostasis-related proteins, which are differentially expressed. Recent studies demonstrate that hESCs have increased proteasome activity in comparison to their differentiated counterparts. This increased proteasome activity was found to be essential for hESC identity and their differentiation into neural cells. Elevated expression of PSMD11, a subunit of the 19S proteasome, is required for the enhanced proteasome activity in hESCs. In addition, ectopic expression of rpn-6 (C. elegans orthologue of PSMD11) was sufficient to confer proteotoxic stress resistance and extend lifespan in this model organism. In this study we show that PSME4, a distinct form of proteasome activator, resembles the high expression levels of PSMD11 in hESCs. However, unlike PSMD11, this protein was found to be III dispensable for stem cell function and organismal longevity under normal growth conditions. Besides the proteasome, we also sought to understand how the chaperome regulates immortality and aging. The human chaperome is formed by 332 chaperones and cochaperones that regulate the folding and function of proteins. We show that human pluripotent stem cells exhibit dramatic differences in expression of molecular chaperones. In particular, we observed much higher expression and increased assembly of the TRiC/CCT complex, a chaperonin that facilitates the folding of 10% of the proteome. This highly conserved molecular machine forms a~1MDa complex in the cytosol of eukaryotic cells, consisting of two back-to-back rings, each containing seven to nine subunits of ~60 kDa. TRiC/CCT is responsible for the folding of many essential proteins, including components of the cytoskeleton such as actin and tubulin or cell cycle regulators, and its dysregulation is associated with a myriad of diseases including cancer and neuropathy. We found that ectopic expression of a single subunit (CCT8) is sufficient to increase TRiC/CCT assembly. Moreover, enhanced TRiC/CCT assembly is required for the striking ability of pluripotent stem cells to maintain proteostasis of aggregation- prone huntingtin (HTT), the mutant protein underlying Huntington’s disease (HD). Since the levels of CCT subunits are further decreased in somatic tissues during organismal aging, we examined whether modulation of CCT8 can delay the aging process and proteostasis dysfunction by using C. elegans as a model organism. Notably, upregulation of CCT8 levels in somatic tissues triggers TRiC/CCT assembly and extends organismal lifespan particularly under proteotoxic conditions. Ectopic expression of CCT8 also ameliorates the age-associated demise of proteostasis and corrects proteostatic deficiencies in worm models of Huntington’s disease. Our results suggest proteostasis is a common principle that links organismal longevity with hESC immortality

Mediation of organismal aging and somatic proteostasis by the germline

Experimental interventions that reduce reproduction cause an extension in lifespan. In invertebrates, such as Caenorhabditis elegans, the aging of the soma is regulated by signals from the germline. Indeed, ablation of germ cells significantly extends lifespan. Notably, germline-deficient animals exhibit heightened resistance to proteotoxic stress. This phenotype correlates with increased potential of intracellular clearance mechanisms such as the proteasome and autophagy in somatic tissues. Here we review the molecular mechanisms by which signals from the germline regulate lifespan in C. elegans with special emphasis on clearance mechanisms

Mechanisms of protein homeostasis (proteostasis) maintain stem cell identity in mammalian pluripotent stem cells

Protein homeostasis, or proteostasis, is essential for cell function, development, and organismal viability. The composition of the proteome is adjusted to the specific requirements of a particular cell type and status. Moreover, multiple metabolic and environmental conditions challenge the integrity of the proteome. To maintain the quality of the proteome, the proteostasis network monitors proteins from their synthesis through their degradation. Whereas somatic stem cells lose their ability to maintain proteostasis with age, immortal pluripotent stem cells exhibit a stringent proteostasis network associated with their biological function and intrinsic characteristics. Moreover, growing evidence indicates that enhanced proteostasis mechanisms play a central role in immortality and cell fate decisions of pluripotent stem cells. Here, we will review new insights into the melding fields of proteostasis and pluripotency and their implications for the understanding of organismal development and survival

Somatic increase of CCT8 mimics proteostasis of human pluripotent stem cells and extends C-elegans lifespan

Human embryonic stem cells can replicate indefinitely while maintaining their undifferentiated state and, therefore, are immortal in culture. This capacity may demand avoidance of any imbalance in protein homeostasis (proteostasis) that would otherwise compromise stem cell identity. Here we show that human pluripotent stem cells exhibit enhanced assembly of the TRiC/CCT complex, a chaperonin that facilitates the folding of 10% of the proteome. We find that ectopic expression of a single subunit (CCT8) is sufficient to increase TRiC/CCT assembly. Moreover, increased TRiC/CCTcomplex is required to avoid aggregation of mutant Huntingtin protein. We further show that increased expression of CCT8 in somatic tissues extends Caenorhabditis elegans lifespan in a TRiC/CCT-dependent manner. Ectopic expression of CCT8 also ameliorates the age-associated demise of proteostasis and corrects proteostatic deficiencies in worm models of Huntington's disease. Our results suggest proteostasis is a common principle that links organismal longevity with hESC immortality