Chaperone networks are required for the shearing and generation of transmissible propagons from pre-existing prion aggregates. However, other cellular networks needed for maintaining yeast prions are largely uncharacterized. Here, we establish a novel role for actin networks in prion maintenance. The [PIN + ] prion, also known as [RNQ + ], exists as stable variants dependent upon the chaperone machinery for the transmission of propagons to daughter cells during cell division and cytoplasmic transfer. Loss of the Hsp104 molecular chaperone leads to the growth of prion particles until they are too large to be transmitted. Here, we isolated a unique [PIN + ] variant, which is unstable in actin mutants. This prion loss is observed over many generations, and coincides with the detection of both high molecular weight species of Rnq1 and large visible aggregates that are asymmetrically retained during cell division. Our data suggest that the irregular actin networks found in these mutants may influence propagon number by slowly permitting aggregate growth over time, resulting in the generation of nontransmissible large aggregates. Thus, we show the potential contribution of cytoskeletal networks in the transmission of prion propagons, which parallels models that have been proposed for cell-to-cell transmission of small amyloids in neurodegenerative protein aggregation diseases.

Sup35p is an essential protein in yeast that functions in complex with Sup45p for efficient translation termination. Although some may argue that this function is the only important attribute of Sup35p, there are two additional known facets of Sup35p's biology that may provide equally important functions for yeast; both of which involve various strategies for coping with stress. Recently, the N‐terminal and middle regions (NM) of Sup35p, which are not required for translation termination function, have been found to provide stress‐sensing abilities and facilitate the phase separation of Sup35p into biomolecular condensates in response to transient stress. Interestingly, the same NM domain is also required for Sup35p to misfold and enter into aggregates associated with the [PSI+] prion. Here, we review these three different states or “faces” of Sup35p. For each, we compare the functionality and necessity of different Sup35p domains, including the role these domains play in facilitating interactions with important protein partners, and discuss the potential ramifications that each state affords yeast cells under varying environmental conditions.

Yeast prions are self-perpetuating misfolded proteins that are infectious. In yeast, [PSI+] is the prion form of the Sup35 protein. While the study of [PSI+] has revealed important cellular mechanisms that contribute to prion propagation, the underlying cellular factors that influence prion formation are not well understood. Prion formation has been described as a multi-step process involving both the initial nucleation and growth of aggregates, followed by the subsequent transmission of prion particles to daughter cells. Prior evidence suggests that actin plays a role in this multi-step process, but actin’s precise role is unclear. Here, we investigate how actin influences the cell’s ability to manage newly formed visible aggregates and how actin influences the transmission of newly formed aggregates to future generations. At early steps, using 3D time-lapse microscopy, several actin mutants, and Markov modeling, we find that the movement of newly formed aggregates is random and actin independent. At later steps, our prion induction studies provide evidence that the transmission of newly formed prion particles to daughter cells is limited by the actin cytoskeletal network. We suspect that this limitation is because actin is used to possibly retain prion particles in the mother cell.

Prions are misfolded, aggregated, infectious proteins found in a range of organisms from mammals to bacteria. In mammals, prion formation is difficult to study because misfolding and aggregation take place prior to symptom presentation. The study of the yeast prion [PSI+], which is the misfolded infectious form of Sup35p, provides a tractable system to monitor prion formation in real time. Recently, we showed that the de novo formation of prion aggregates begins with the appearance of highly mobile cytoplasmic foci, called early foci, which assemble into larger ring or dot structures. We also observed SDS-resistant oligomers during formation, and lysates containing newly formed oligomers can convert [psi−] cells to the [PSI+] state, suggesting that these oligomers have infectious potential. Here, we further characterize two aspects of prion formation: spatial sequestration of early foci and oligomerization of endogenous Sup35p. Our data provides important insights into the process of prion formation and explores the minimal oligomer requirement for infectivity.