Introduction
The prion-folding states orchestrated by chaperone systems in protein
homeostasis are closely related to prion diseases, such as transmissible
spongiform encephalopathy (TSE) [1-3]. Previous studies have
revealed that prion transmission is a universal phenomenon, which shows
intriguingly similar self-assembly behavior and heredity stability fromE. coli and yeast to mammals [4-6].However, cells in
different organisms exhibit distinct responses and regulatory mechanisms
for prion assembly [7]. Investigations on the heterologous
expression of a prion protein in a foreign host should allow one to gain
insight into the prion self-assembly mechanism.
S. cerevisiae Sup35 protein is an ideal model for examining prion
self-assembly and genetic stability from parental cells to filial cells.
In its normal monomer state, Sup35 serves as a translation termination
release factor, dictated by the C-terminal [8,9]. In contrast, the
N-terminal, comprising relatively conserved amino acids, serves as the
core functional domain for amyloid formation [10]. Sup35-NM, which
was constructed using the N-terminal and middle (M) domain, is typically
chosen for in vitro self-assembly studies. Transforming in
vitro -prepared amyloid fibrils into non-prion state yeast cells can
effectively induce prion state phenotypes, and these phenotypes possess
genetic stability from one generation to the next [11,12].Meanwhile,
Sup35 overexpression in yeast causes the prion state phenotype
[PSI +] [13], which limits the
investigation of prion transmission in the in vivo state by
homologous overexpression. HSP104 has been suggested as a contributing
factor in the development of prion states in
[PSI +] [14,15], but the process by
which exhaustive genetic and self-assembly stability is maintained in
yeast cells has remained unclear.
Many conclusions about the in vitro prion self-assembly mechanism
have been drawn from prokaryotic purified proteins [16-18].
Examinations of Sup35 heterologous expression have primarily focused on
two aspects. For one, GPI-anchored Sup35 in mammalian cells can form
aggregates, which are different from the fibrils assembled in
vitro [19,20]. This suggests that heterologous expression is
feasible and can contribute to the understanding of the prion
self-assembly mechanism. In addition, studies examining Sup35 and
Sup35-NM self-assembly have been conducted in E. coli . Sup35-NM
fusion expression with green fluorescent protein in E. colishowed bright foci and amyloid-like structures [21]. Several lines
of evidence have shown that overexpressed Sup35 could be purified in
soluble states in E. coli [22,23]. In contrast, Sup35 from
yeast formed high molecular weight aggregates. These findings suggest
that Sup35 may exist in the cytoplasm of E. coli cells as
monomers. The cellular components responsible for maintaining the
monomer state for Sup35 in heterologous E. coli are not fully
understood. Furthermore, whether Sup35 monomer constitutes the sole
surviving state in E. coli remains unclear. Answering these
questions will provide a solid foundation for achieving prion
self-assembly in a controlled fashion and can offer deeper insight into
prion in vivo heredity and infectivity.