Date Published: July 13, 2017
Publisher: John Wiley and Sons Inc.
Author(s): Zhonghe Ke, Pramit Mallik, Adam B. Johnson, Facundo Luna, Eviatar Nevo, Zhengdong D. Zhang, Vadim N. Gladyshev, Andrei Seluanov, Vera Gorbunova.
Whether errors in protein synthesis play a role in aging has been a subject of intense debate. It has been suggested that rare mistakes in protein synthesis in young organisms may result in errors in the protein synthesis machinery, eventually leading to an increasing cascade of errors as organisms age. Studies that followed generally failed to identify a dramatic increase in translation errors with aging. However, whether translation fidelity plays a role in aging remained an open question. To address this issue, we examined the relationship between translation fidelity and maximum lifespan across 17 rodent species with diverse lifespans. To measure translation fidelity, we utilized sensitive luciferase‐based reporter constructs with mutations in an amino acid residue critical to luciferase activity, wherein misincorporation of amino acids at this mutated codon re‐activated the luciferase. The frequency of amino acid misincorporation at the first and second codon positions showed strong negative correlation with maximum lifespan. This correlation remained significant after phylogenetic correction, indicating that translation fidelity coevolves with longevity. These results give new life to the role of protein synthesis errors in aging: Although the error rate may not significantly change with age, the basal rate of translation errors is important in defining lifespan across mammals.
The error catastrophe theory of aging was proposed by Orgel in the 1960s (Orgel, 1963, 1970, 1973). According to this model, the aging process results from errors in mRNA translation that reduce the fidelity of the protein‐translating enzymes leading to increasingly inaccurate protein synthesis, terminating in functional decline, and, ultimately, the death of the organism (Srivastava & Busbee, 2003). This theory, for the first time, proposed that translation fidelity plays a major role in aging.
To measure the fidelity of protein synthesis, we used sensitive luciferase‐based reporters that we previously developed (Azpurua et al., 2013) (Fig. 2A). In these reporters, the firefly luciferase gene is mutated at position K529, the residue essential for luciferase activity (Kramer & Farabaugh, 2007). The mutations include nucleotide substitutions at the first (K529E), second (K529I), and third (K529N) codon positions and also a substitution to a stop codon (TGA) at the amino acid position 81. Amino acid misincorporation at these positions results in reactivation of the luciferase gene and is scored using a luminometer. Primary, low passage, rodent fibroblasts from 17 species with diverse lifespans from our collection (Seluanov et al., 2008, 2010; Patrick et al., 2016) were cotransfected with the luciferase reporters and Renilla luciferase as a transfection control. Three independent (derived from different animals) cell lines were assayed for each species, and each cell line was tested in triplicate.
This work was supported by grants from US National Institutes of Health to VG, AS, ZZ, and VNG; Life Extension Foundation to VG and AS, Consejo Nacional de Investigaciones Científicas y Técnicas to FL, and NY Stem predoctoral fellowship to ZK.