Structural Elements of ‘Anti‐prion J protein 1’ (Apj1) Required for Efficient Curing of the [PSI+] Prion by Hsp104 Overexpression

Abstract

Prions are self‐propagating aggregates of misfolded protein that often form ordered amyloids. Amyloids are associated with a variety of human neurodegenerative diseases, including but not limited to Alzheimer’s disease, Huntington’s disease, and Parkinson’s disease. Amyloid‐forming prions are also present in Saccharomyces cerevisiae, where they are propagated through populations by a fragmentation mechanism that requires three chaperone proteins: the disaggregase Hsp104, the Hsp70 Ssa, and the J‐protein Sis1. When Hsp104 is ectopically overexpressed, however, the prion [PSI+] is specifically eliminated from cell populations in a Sis1‐dependent manner. We previously determined that the J‐protein Apj1 is also required for efficient elimination, or curing, of the prion variants [PSI+]STR and [PSI+]Sc4 by Hsp104 overexpression. Additionally, we showed that Apj1 and Sis1 have overlapping functions in the Hsp104‐mediated curing process, as overexpressing one in the absence of a functional version of the other restores curing. A construct bearing the first 161 residues of Apj1 is sufficient to restore efficient curing in Δapj1cells. Residues 1‐161 can be subdivided based on sequence features and homology into: a J‐domain, a polyglutamine track with an imperfect QA repeat region, a region bearing homology to the G/F region of Sis1, and finally a region rich in glycine, serine, phenylalanine, and asparagine which resembles the amino acid content of many prion‐forming domains. Various constructs produced by mutating and truncating regions of Apj1‐161 were ectopically expressed in Δapj1 cells to determine which regions, if any, were sufficient to replace Apj1 in Hsp104‐mediated curing. We found that constructs containing the polyQ region restored efficient curing and those lacking the polyQ region did not support efficient curing, indicating the importance of the polyQ region in the curing process. The Sis1 homologous region and the SFN rich region proved nonessential for Hsp104‐mediated curing. Ongoing experiments are being conducted to determine which regions of Apj1‐161 are required to replace the function of Sis1 in Hsp104‐mediated curing.