Rapid selection of cyclic peptides that reduce α-synuclein toxicity in yeast and animal models

Abstract

Phage display has demonstrated the utility of cyclic peptides as general protein ligands, but cannot access proteins inside eukaryotic cells. Expanding a novel chemical genetics tool, we describe the first expressed library of head-to-tail cyclic peptides in yeast (Saccharomyces cerevisiae). We applied the library to selections in a yeast model of α-synuclein toxicity that recapitulates much of the cellular pathology of Parkinson’s disease. From a pool of five million transformants, we isolated two related cyclic peptide constructs which specifically reduce the toxicity of human α-synuclein. These expressed cyclic peptide constructs also prevent dopaminergic neuron loss in an established Caenorhabditis elegans Parkinson’s model. This work highlights the speed and efficiency of using libraries of expressed cyclic peptides for forward chemical genetics in cellular models of human disease.

Compounds from an unbiased chemical screen reverse both ER-to-Golgi trafficking defects and mitochondrial dysfunction in Parkinson’s disease models

“…The Rab proteins are largely ineffectual in protecting HiTox cells on their own ). They do so only when combined with genetic suppressors of different functional classes Kritzer et al, 2009). We found that, unlike the Rab proteins, all four bioactive compounds restored -syn localization to the plasma membrane on their own in HiTox cells (Fig.…”

“…The IntTox strain also can be suppressed by single genetic modifiers (Yeger-Lotem et al, 2009). By contrast, the HiTox -syn strain, which only expresses ~50% more -syn, requires two genetic suppressors acting in separate pathways to rescue growth Kritzer et al, 2009). Transcriptional profiling revealed that, in this strain, in addition to ER-to-Golgi trafficking defects, mitochondrial dysfunction is an early pathological phenotype.…”

“…In various experiments using ELISA assays, the expression level of -syn in HiTox cells was 1.4-to 2.5-fold higher than the -syn expression level in the IntTox cells. One striking difference between these two strains is that most of the genetic modifiers that strongly suppress -syn toxicity in the IntTox strain do not effectively suppress toxicity in the HiTox strain on their own Kritzer et al, 2009). Instead, co-expression of two suppressors from diverse pathways is required to restore viability in the HiTox strain.…”

BacHBerry: BACterial Hosts for production of Bioactive phenolics from bERRY fruits

“…Consequently, polyphenol-based therapies represent an underexplored strategy to minimize the huge social and economic impact of NDs. The high degree of evolutionary conservation of fundamental biological processes among eukaryotes has established baker's yeast as a validated model organism to decipher the intricacies of human pathologies, particularly NDs, to identify molecular targets amenable to therapeutic intervention as well as lead molecules with health-promoting potential (Kritzer et al 2009;Su et al 2010;Tardiff et al 2012;Menezes et al 2015). BacHBerry aimed at identifying phenolic bioactives from harnessing the diversity of phenolics in selected berry germplasm from cultivated, wild and underutilized species of berries.…”

Methods for the Creation of Cyclic Peptide Libraries for Use in Lead Discovery

“…11,18,19 The latter uses a selectively randomized library of splitinteins for the production of genetically encoded, backbone cyclized peptide libraries. Semisynthetic libraries are able to incorporate nonpeptidogenic amino acids while retaining ribosomal synthesis and an associated genetic tag.…”