C. elegans Models to Study the Propagation of Prions and Prion-Like Proteins

Abstract

A hallmark common to many age-related neurodegenerative diseases, such as Alzheimer's disease (AD), Parkinson's disease (PD), and amyotrophic lateral sclerosis (ALS), is that patients develop proteinaceous deposits in their central nervous system (CNS). The progressive spreading of these inclusions from initially affected sites to interconnected brain areas is reminiscent of the behavior of bona fide prions in transmissible spongiform encephalopathies (TSEs), hence the term prion-like proteins has been coined. Despite intensive research, the exact mechanisms that facilitate the spreading of protein aggregation between cells, and the associated loss of neurons, remain poorly understood. As population demographics in many countries continue to shift to higher life expectancy, the incidence of neurodegenerative diseases is also rising. This represents a major challenge for healthcare systems and patients' families, since patients require extensive support over several years and there is still no therapy to cure or stop these diseases. The model organism Caenorhabditis elegans offers unique opportunities to accelerate research and drug development due to its genetic amenability, its transparency, and the high degree of conservation of molecular pathways. Here, we will review how recent studies that utilize this soil dwelling nematode have proceeded to investigate the propagation and intercellular transmission of prions and prion-like proteins and discuss their relevance by comparing their findings to observations in other model systems and patients.

Keywords: C. elegans; intercellular spreading; neurodegenerative diseases; prion-like propagation; prions.

Figure 1

Overview of the currently established C. elegans models for prion-like propagation. The three different types of model systems and their working principles are illustrated using α-Syn and TTR as examples. Further models for other prion-like proteins are described in the text and listed in Table 1. (A) Schematic overview of C. elegans with the locations of the longitudinal and cross-sections, which are shown in more detail in panel B–D. (B–D) Longitudinal (B) or cross-sections (C,D) on the left show exemplary cells of the donor & recipient tissues within the worm. On the right-side, expression and transmission of the respective prion-like protein is shown schematically. Thick edges, as seen in pharynx and body wall muscle indicate presence of a tissue border (basement membrane). Large symbols represent construct that is expressed in the respective cell, while smaller symbols represent transmitted proteins and proteins formed after transmission. (B) Representation of C. elegans models using the bimolecular fluorescence complementation (BiFC) method to study the spreading of α-Syn. Here, α-Syn expressed in either the pharynx or pharyngeal neurons (or different neuronal subsets) is coupled with separate fluorophore fragments (e.g., Venus or GFP), which only exhibit fluorescence upon excitation, when they complement to a functional protein. Close proximity of the fragments, as in α-Syn aggregation, is required for complementation to happen. Thus, fluorescence observed in either cell indicates intercellular exchange of the α-Syn, as well as close proximity. (C) Representation of C. elegans models using fluorescent proteins (here mRFP) fused to α-Syn to track transmission from the donor tissue, in which expression takes place (here the body wall muscle or dopaminergic neurons), to surrounding recipient tissues (here hypodermis). (D) Representation of C. elegans models in which transthyretin (TTR) is expressed without any tag in the donor tissue (here the body wall muscle). Transmission into recipient tissues, such as the coelomocytes, is then studied by the addition of a compound which binds TTR tetramers. This binding can then be visualized by fluorescence microscopy. Non-cell autonomous effects of TTR secretion and accumulation in or near sensory neurons, can be indirectly inferred by behavioral assays reporting on sensory neuron function.

Figure 2

Schematic overview of the cellular pathways involved in the propagation of prion-like aggregates within and between C. elegans tissues. In the donor tissue, native monomers may spontaneously or due to cellular stress acquire a pathological β-sheet rich conformation, which in turn can seed the misfolding of cognate monomers and thus grow into larger fibrils that eventually deposit into large cytoplasmic inclusions. While the exact nature of the toxic protein species is still unclear, the process of aggregate formation has been clearly linked to cell autonomous toxicity. The cellular chaperone network attempts to dissociate protein aggregates, resulting in the generation of additional seeds that may promote prion-like propagation. Misfolded and aggregated protein species are recognized by the autophagy machinery and subsequently either targeted for lysosomal degradation or removal via exocytosis. Aggregated material expelled from the donor tissue can end up in extracellular deposits that can damage adjacent tissues non-cell autonomously, or it can be taken up via endocytosis into the surrounding tissue. From the endocytic compartment of the recipient cell, seeds may enter the cytosol after vesicle rupture. Once in the recipient cytoplasm, misfolding of cognate native monomers can be templated by the seeds, which also confers non-cell autonomous toxicity. Factors known to impact prion-like propagation and toxicity, are for example ageing (AGE) and the impairment of insulin signaling (INS), the effects of which are indicated in the figure at relevant locations.