Physiological and environmental control of yeast prions

Abstract

Prions are self-perpetuating protein isoforms that cause fatal and incurable neurodegenerative disease in mammals. Recent evidence indicates that a majority of human proteins involved in amyloid and neural inclusion disorders possess at least some prion properties. In lower eukaryotes, such as yeast, prions act as epigenetic elements, which increase phenotypic diversity by altering a range of cellular processes. While some yeast prions are clearly pathogenic, it is also postulated that prion formation could be beneficial in variable environmental conditions. Yeast and mammalian prions have similar molecular properties. Crucial cellular factors and conditions influencing prion formation and propagation were uncovered in the yeast models. Stress-related chaperones, protein quality control deposits, degradation pathways, and cytoskeletal networks control prion formation and propagation in yeast. Environmental stresses trigger prion formation and loss, supposedly acting via influencing intracellular concentrations of the prion-inducing proteins, and/or by localizing prionogenic proteins to the prion induction sites via heterologous ancillary helpers. Physiological and environmental modulation of yeast prions points to new opportunities for pharmacological intervention and/or prophylactic measures targeting general cellular systems rather than the properties of individual amyloids and prions.

Keywords: amyloid; chaperone; cytoskeleton; heat shock; quality control; ubiquitin.

Figure 1. Detection of prion [PSI⁺] by the nonsense-suppression assay

In a [psi⁻] ade1-14 reporter strain, stop codon UGA, introduced in the middle of ADE1 gene, is normally recognized by the translation termination complex including release factor Sup35. This results in termination of translation, synthesis of truncated Ade1 protein, inability of cells to grow on the medium lacking adenine (–Ade) and accumulation of red color on complete YPD medium. In the [PSI⁺] strain, Sup35 is present in the partially inactive prion form and termination is inefficient, resulting in the synthesis of complete Ade1 protein, growth on –Ade medium and white color on YPD medium. Other phenotypes associated with [PSI⁺] presence include: aggregation of Sup35-GFP, distribution of Sup35 into the pellet fraction after centrifugation of cell lysates, presence of SDS-resistant polymers (see Liebman & Chernoff, 2012).

Figure 2. A model of prion destabilization by alteration of the chaperone balance during stress

In the exponentially growing yeast cells, Hsp104 is present at a low level. It is rapidly accumulated during the first 30 min of heat shock and only moderately rises further during next 3.5 hrs at high temperature. In contrast, Hsp70-Ssa is present at the considerably higher background levels in exponential cells, compared to Hsp104. As a result, its levels rise slowly during heat shock, reaching maximum only by about 4 hrs of incubation at high temperature. Efficient fragmentation of Sup35 prion aggregates occurs when Hsp70-Ssa binds to aggregates and recruits Hsp104 to break prion polymers into prion fragments, propagons, inheritable by a daughter cell. When Hsp104 is present in an excess, it binds prion polymers on its own in the absence of Hsp70-Ssa but is not able to efficiently fragment them. We propose that the same occurs after short-term exposure to heat shock, as confirmed by increased size of Sup35 aggregates. As a result, large aggregates become trapped in the mother cell after a resumption of cell division, possibly in association with the quality control deposits of heat-damaged proteins. This apparently leads to prion clearance from a daughter cell, detected in the experiment. After longer incubation at high temperature, the balance between Hsp104 and Hsp70-Ssa is partly restored. This coincides with the prion recovery, suggesting that normal fragmentation and propagation of aggregates is resumed due to restoration of the chaperone balance. Images show colonies of a yeast strain bearing a weak prion variant, that are plated after various exposures to 39°C. Red color indicates prion loss. For details, see (Chernova, et al., 2011, Newnam, et al., 2011).

Figure 3. Preexisting Q/N-rich aggregates promote conversion of soluble heterological protein into a prion form

(A) Cross-seeding model: Q/N rich aggregate (green) serves as a nucleation factor for initial aggregation of prion protein (red). (B) Time-lapse confocal microscopy images of cells expressing Q/N rich protein Lsb2-GFP and prion protein Sup35NM-dsRED (Chernova, et al., 2011). Small aggregates of Sup35 fuse and grow into large aggregates in dynamic association with Lsb2. Aggregates of Lsb2 seed conversion of Sup35 into prion [PSI⁺].

Figure 4. Model for the role of Lsb2 as a stress-dependent prion auxillary factor

Under normal conditions, levels of the Q/N rich aggregation-prone cytoskeleton-associated protein Lsb2 in the cell are low and are tightly regulated by ubiquitinproteasome system. When level of Lsb2 is increased, protein become ubiquitinated (higher molecular weight bands) and quickly degraded. Short-term rise of the temperature to 39°C leads to strong increase in the Lsb2 protein levels, that may trigger the accumulation of misfolded Sup35 at the cytoskeleton-associated cortical locations. In the [PSI⁺] cells, this partially protects [PSI⁺] from uncontrolled agglomeration and elimination. In a small fraction of [psi⁻] cells, Lsb2 could seed conversion of accumulated Sup35 into the prion form [PSI⁺]. The proteolytically stable prion aggregates escape the quality control compartments, while Lsb2 aggregates are recycled and/or degraded.