Cell-penetrable, protease-resistant αA-crystallin-derived peptide chaperone triggers cell stress response pathways and alleviates β-amyloid engendered toxicity in Caenorhabditis elegans

Abstract

Purpose : Mini-chaperones derived from αA-crystallin, with their inherent anti-aggregation and anti-apoptotic properties, have therapeutic properties. A second generation mini-chaperone peptide, having cell-penetrating and protease-resistant properties (CPPRMC), was synthesized using L- and D- amino acids. This study aimed to evaluate the therapeutic efficacy of CPPRMC and unravel the mechanism of peptide action using cell culture and C. elegans model systems.

Methods : The anti-oxidant and anti-apoptotic properties of the CPPRMC was evaluated in sodium iodate-treated ARPE-19 cells. The effect of peptide treatment (50 μg every two days along with E. coli OP50 food source) on the wild-type (N2) worm lifespan was tested at 20^oC. Tolerance to chemical-stimulated oxidative stress (50 mM Paraquat, 50 µM Juglone, and 40 mM glucose) was evaluated using peptide-fed N2 worms (Day-5). The CPPRMC's ability to attenuate beta-amyloid aggregation-induced paralysis and survival was tested in the CL4176 strain expressing β-amyloid peptide. RNA sequencing and high-throughput untargeted metabolic profiling (polar and non-polar) were also performed to compare differentially expressed genes and metabolites across experimental conditions.

Results : The Sodium iodate-induced (1 mM) oxidative stress and cytotoxicity were suppressed entirely in CPPRMC treated (5 μM) ARPE-19 cells. Peptide treatment increased the lifespan of N2 worms by 18%. N2 worms pretreated with 50 µg CPPRMC for 24h at 20^oC and then subjected to thermal stress at 35^oC showed a 43% increase in survival as equated to vehicle control. Peptide treatment delayed AD-related paralysis and extended worm's survival (17%) after temperature up-shift in CL4176. RNA-seq analysis showed the upregulation of genes responsible for lifespan increase, and the polar and non-polar metabolic profiling revealed altered metabolites involved in the lifespan.

Conclusions : The increased worm lifespan and alleviation of Aβ-induced toxicity up on CPPRMC treatment are likely mediated by the activation of molecules involved in stress response pathways. Our findings signify the therapeutic prospective of CPPRMC against aging and age-concerned AD.

This is a 2021 ARVO Annual Meeting abstract.