Protein Homeostasis Networks and the Use of Yeast to Guide Interventions in Alzheimer's Disease

Abstract

Alzheimer's Disease (AD) is a progressive multifactorial age-related neurodegenerative disorder that causes the majority of deaths due to dementia in the elderly. Although various risk factors have been found to be associated with AD progression, the cause of the disease is still unresolved. The loss of proteostasis is one of the major causes of AD: it is evident by aggregation of misfolded proteins, lipid homeostasis disruption, accumulation of autophagic vesicles, and oxidative damage during the disease progression. Different models have been developed to study AD, one of which is a yeast model. Yeasts are simple unicellular eukaryotic cells that have provided great insights into human cell biology. Various yeast models, including unmodified and genetically modified yeasts, have been established for studying AD and have provided significant amount of information on AD pathology and potential interventions. The conservation of various human biological processes, including signal transduction, energy metabolism, protein homeostasis, stress responses, oxidative phosphorylation, vesicle trafficking, apoptosis, endocytosis, and ageing, renders yeast a fascinating, powerful model for AD. In addition, the easy manipulation of the yeast genome and availability of methods to evaluate yeast cells rapidly in high throughput technological platforms strengthen the rationale of using yeast as a model. This review focuses on the description of the proteostasis network in yeast and its comparison with the human proteostasis network. It further elaborates on the AD-associated proteostasis failure and applications of the yeast proteostasis network to understand AD pathology and its potential to guide interventions against AD.

Keywords: Alzheimer’ disease; autophagy; proteostasis; ubiquitin proteasome system; unfolded protein response; yeast.

Figure 1

Risk factors/loci and associated cellular processes identified by genome wide association studies (GWAS) involved in Alzheimer’s Disease (AD) converge to the proteostasis network. APP, amyloid precursor protein; PSEN1, presenilin 1; PSEN2, presenilin 2; ADAM10, ADAM metallopeptidase domain 10; CLU, clusterin; CR1, complement C3b/C4b receptor 1; ABCA7, ATP binding cassette subfamily A member 7; CD33, CD33 molecule; MS4A, membrane spanning 4-domains A; PICALM, phosphatidylinositol binding clathrin assembly protein; CD2AP, CD2 associated protein; EPHA1, EPH receptor A1; BIN1, bridging integrator 1; APOε4, apolipoprotein E4; SORL1, sortilin related receptor 1. Various risk loci have been identified by GWAS to be associated with AD. Some of these risk loci are clustered to important cellular processes in this figure to depict their relationship with the proteostasis network, highlighting its significance in AD pathology. Previous studies suggest an important role of the proteostasis network in cellular processes associated with AD, including amyloid formation pathway, immune response mechanisms, vesicle trafficking, endocytosis, and lipid metabolism.

Figure 2

Proteostasis network involves protein synthesis, protein folding, heat shock response, unfolded protein response, ubiquitin proteasome system, and autophagy. Green arrow, protective; Red arrow, toxic; HSF1, heat shock factor 1; HSE, heat shock elements; UPRE, unfolded protein response elements. The proteostasis network involves protein synthesis machinery, protein folding, protein quality control, protein transport, and the overall turnover of proteins inside a cell. The presence of misfolded proteins in the cytosol trigger HSF1 activation, which activates promoters with HSE and results in expression of heat shock proteins that are either involved in protein disaggregation and refolding or clearance of the unwanted proteins through protein quality control mechanisms. Similarly, the presence of aberrations in the ER lumen causing unfolded protein formation leads to activation of the unfolded protein response and downstream activation of the protein quality control system.

Figure 3

Schematic diagram showing unfolded protein response in yeast cells. Bip, binding immunoglobulin protein; ER, endoplasmic reticulum; Ire1, inositol requiring element 1; HAC1, Homologous to ATF/CREB1; Rlg1p, tRNA ligase protein; UPRE, unfolded protein response elements. The presence of unfolded/misfolded proteins in the ER lumen is recognized by the yeast Ire1 sensor protein that activates the specific cytosolic endonuclease activity cleaving the inhibitory intron of HAC1 mRNA and rendering HAC1 mRNA translationally active. Hac1p protein translates and translocates to the nucleus, where it recognizes the characteristic cis-acting elements in the promoter regions of certain genes referred to as UPRE and increases expression of genes under control of such promoters. The process is referred to as the UPR.

Figure 4

Schematic diagram showing degradation of substrate protein by the UPS and fates of defective proteasomal system. Ub, ubiquitin; ATP, adenosine triphosphate. The first step in ubiquitin proteasome system requires the activated E1 activating enzyme to activate the ubiquitin. Following the activation of ubiquitin, E1 enzyme transfers the activated ubiquitin molecule to E2 conjugating enzyme. At the same time, E3 ligating enzyme conjugates with the substrate protein, which is then conjugated with ubiquitin bound to E2 enzyme. The chain of ubiquitin is elongated by multiple cycles of ubiquitination. The ubiquitinated proteins are then recognized and degraded by the 26S proteasome, while the failure to degrade the polyubiquitinated proteins due to defective proteasome may result in protein aggregation, abundance of polyubiquitinated proteins, and impaired protein turnover. The defective proteasome may get recycled through proteaphagy.

Figure 5

Schematic diagram of autophagosome formation and delivery of embedded cargo to the vacuole. Arrow, activates; Blunt arrow, blocks/inhibits; Atg, autophagy related; PI3K, phosphatidyl inositol 3 kinase; PE, phosphatidylethanolamine. Autophagosome formation is completed in four steps starting from the activation of Atg13 and formation of Atg1 kinase complex at the pre-phagosome assembly site. The second step involves the initiation of the phagophore formation by recruiting Atg9 protein. The phagophore nucleation occurs following Atg9 conjugation, where complexation with phosphatidyl inositol 3 kinase complex I occurs. The expansion of the double walled phagophore occurs embedding the cytosolic cargo that will ultimately give rise to autophagosome. The mature autophagosome then delivers the embedded cargo to the vacuole for degradation.

Figure 6

Different routes of delivery of cytosolic materials into the vacuole of yeast highlighting the fusion of autophagosomes with vacuole and energy-dependent role of V-ATPase in maintaining the pH of vacuoles. In the figure, the fusion of the autophagosome and the lysosomal membrane releases the autophagosomal cargo to the acidic lumen of the vacuole in yeast where these cargoes are degraded with the help of vacuole resident pH dependent cathepsins, hydrolases, and lipases. The pH of the vacuolar lumen is tightly regulated by an ATP dependent proton pump, the V-ATPase. Apart from the autophagosome mediated delivery of cargo inside vacuoles, endocytic vesicles and Golgi bodies carrying defective proteins are also delivered to the vacuole for degradation. Lysosomes may also degrade proteins and help protein turnover by direct engulfment of some cytosolic proteins, the process referred to as microautophagy.

Figure 7

Some possible cell pathways that converge to yeast autophagy in response to various stimuli, including nutrient sensing, calcium homeostasis, oxidative stress, misfolded proteins, and UPR. Nutrient supplementation in a eukaryotic cell activates RAS signaling activating protein kinase activity leading to the activation of mTOR, whereas nutrient deficiency activates AMPK/Snf1 via sensing AMP/ATP ratio changes. Sir2/SIRT1 gets activated by increasing NAD⁺ in an energy-deficient environment and activates several downstream targets via deacetylation of its targets. Nutrient starvation may also have role in inefficient protein folding triggering the formation of unfolded proteins and misfolded proteins. The folding aberrations in ER are indicated by altered calcium levels in the cytosol and ER lumen. The increase in cytosolic calcium activates CAMKKβ, an upstream kinase of AMPK. The calcium abundance may also cause calcium overloading of mitochondria and enhance ROS formation. The presence of unfolded protein activates UPR, antioxidant response, MAPK signaling, and FOXO signaling and supports autophagy induction. However, chronic ER stress and JNK signaling may trigger apoptosis activation. The interaction between AMPK/Snf1, FOXO/Hcm1, and SIRT1/Sir2 induces autophagy and cytoprotective processes, whereas activation of mTOR, chronic ER stress, mitochondrial calcium overloading, and ROS may decrease survival possibly via inhibiting autophagy. During metabolic stress, DNA damage and oxidative damage, autophagy may also be activated via p53 activation. Meanwhile, p53 may also trigger apoptosis and cell cycle arrest depending on the cellular environment. Regulation of autophagy is a complex interaction of intracellular cues acting at several different levels. Some mechanisms depicted in the figure in the red boxes are not reported to be present in yeast.