The number and transmission of [PSI] prion seeds (Propagons) in the yeast Saccharomyces cerevisiae

Abstract

Background: Yeast (Saccharomyces cerevisiae) prions are efficiently propagated and the on-going generation and transmission of prion seeds (propagons) to daughter cells during cell division ensures a high degree of mitotic stability. The reversible inhibition of the molecular chaperone Hsp104p by guanidine hydrochloride (GdnHCl) results in cell division-dependent elimination of yeast prions due to a block in propagon generation and the subsequent dilution out of propagons by cell division.

Principal findings: Analysing the kinetics of the GdnHCl-induced elimination of the yeast [PSI+] prion has allowed us to develop novel statistical models that aid our understanding of prion propagation in yeast cells. Here we describe the application of a new stochastic model that allows us to estimate more accurately the mean number of propagons in a [PSI+] cell. To achieve this accuracy we also experimentally determine key cell reproduction parameters and show that the presence of the [PSI+] prion has no impact on these key processes. Additionally, we experimentally determine the proportion of propagons transmitted to a daughter cell and show this reflects the relative cell volume of mother and daughter cells at cell division.

Conclusions: While propagon generation is an ATP-driven process, the partition of propagons to daughter cells occurs by passive transfer via the distribution of cytoplasm. Furthermore, our new estimates of n(0), the number of propagons per cell (500-1000), are some five times higher than our previous estimates and this has important implications for our understanding of the inheritance of the [PSI+] and the spontaneous formation of prion-free cells.

Figure 1. Simulation showing the proportion of [PSI ⁺] cells against time following the addition of 3 mM GdnHCl to a dividing yeast culture.

The data (denoted by X) are simulated by assuming that cells start with an average of n ₀ = 100 propagons per cell. The time that a mother cell takes to divide follows a gamma distribution with a mean of 2 hr and standard deviation 0.28 hr. The total time that a daughter cell takes to divide also follows a gamma distribution, but with means 3 hr and standard deviation 0.35 hr. The probability that a propagon passes to a daughter cell is assumed to be 0.3. Three different models are shown fitted to the data: −·−·−· Model A ; - - - - Model B and − Model C which gives an estimate of π = 0.3 (with standard error 0.02). The value of n ₀ estimated by each model is shown with standard errors in brackets.

Figure 2. A ‘division tree’ diagram showing the growth and division of yeast cells from a single original mother cell.

The original mother cell is labelled “1”. Each time a new bud (daughter) appeared on the original mother cell it was labelled 1.1, 1.2, 1.3 etc…and the time of appearance noted. When these daughter cells produced buds they were labelled 1.1.1, 1.1.2 etc… Two sets of data could be derived: a) the successive time taken for a cell to divide multiple times (mother cell reproduction times), and b) the time taken from emergence as a bud on a mother cell to subsequent first division (daughter cell reproduction times).

Figure 3. Observed proportion of [PSI ⁺] cells and fitted curve of p₊(t).

One set of data for each of the [PSI ⁺] strains (A) YJW512 and (B) YJW679 is shown. The curing parameter estimates for three independent experiments (1, 2, and 3) with 3 mM of GdnHCl are given in the table (C) with the values in brackets being the estimated standard errors for each parameter. Panels A and B represent one experiment while two other sets of data independently generated for these two strains are provided in Figure S1.

Figure 4. The effects of different concentrations of GdnHCl on the kinetics of elimination of the [PSI ⁺] prion from the strain YJW512.

(A) The proportion of cells in the population at a given time is plotted against the expected generation number . For each data set a fitted curve has been added. (B) Curing parameter estimates for different concentrations of GdnHCl. Values in brackets are the estimated standard errors for the parameters.

Figure 5. Proportion of prions passed on to the daughter cell for [PSI ⁺] strains YJW512 (A) and YJW679 (B).

The maximum likelihood estimate of π (with standard error in brackets) shown is based on the assumption the number of prions passed on to a daughter cell follows a binomial distribution, as used in Model C (see .