Abstract
[PSI+], the prion form of the yeast Sup35 protein, results from the structural conversion of Sup35 from a soluble form into an infectious amyloid form. The infectivity of prions is thought to result from chaperone-dependent fiber cleavage that breaks large prion fibers into smaller, inheritable propagons. Like the mammalian prion protein PrP, Sup35 contains an oligopeptide repeat domain. Deletion analysis indicates that the oligopeptide repeat domain is critical for [PSI+] propagation, while a distinct region of the prion domain is responsible for prion nucleation. The PrP oligopeptide repeat domain can substitute for the Sup35 oligopeptide repeat domain in supporting [PSI+] propagation, suggesting a common role for repeats in supporting prion maintenance. However, randomizing the order of the amino acids in the Sup35 prion domain does not block prion formation or propagation, suggesting that amino acid composition is the primary determinant of Sup35's prion propensity. Thus, it is unclear what role the oligopeptide repeats play in [PSI+] propagation: the repeats could simply act as a non-specific spacer separating the prion nucleation domain from the rest of the protein; the repeats could contain specific compositional elements that promote prion propagation; or the repeats, while not essential for prion propagation, might explain some unique features of [PSI+]. Here, we test these three hypotheses and show that the ability of the Sup35 and PrP repeats to support [PSI+] propagation stems from their amino acid composition, not their primary sequences. Furthermore, we demonstrate that compositional requirements for the repeat domain are distinct from those of the nucleation domain, indicating that prion nucleation and propagation are driven by distinct compositional features.
Highlights
The [PSI+] prion of Saccharomyces cerevisiae results from the structural conversion of the Sup35 protein into an infectious amyloid conformation [1,2,3]
The glutamine/ asparagine-rich (Q/N-rich) tract is primarily responsible for prion nucleation and fiber growth, while the remained of the prion-forming domain (PFD) is thought to be primarily involved in prion propagation [8,10,11,12,13]; this region dominated by an oligopeptide repeat domain (ORD) spanning amino acids 40–96
The fusion constructs were named FP21N, FP21C, FP24N, FP24C, FP26N, FP26C, FP27N and FP27C
Summary
The [PSI+] prion of Saccharomyces cerevisiae results from the structural conversion of the Sup protein into an infectious amyloid conformation [1,2,3]. Sup is composed of three functionally and structurally distinct domains (Figure 1): the C-terminal domain (a.a. 254–685) is essential for the translation termination activity, the prion-forming domain (PFD; a.a. 1–114) drives the structural conversion to amyloid, and the highly charged middle domain has no known function other than its ability to stabilize [PSI+] fibers [6,7,8,9]. The Sup PFD is composed of two separate sub-domains thought to have distinct functions (Figure 1A). The glutamine/ asparagine-rich (Q/N-rich) tract (amino acids 1–39) is primarily responsible for prion nucleation and fiber growth, while the remained of the PFD (amino acids 40–114) is thought to be primarily involved in prion propagation [8,10,11,12,13]; this region dominated by an oligopeptide repeat domain (ORD) spanning amino acids 40–96. A large body of evidence supports an essential role for the ORD in prion propagation
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