A Platform for Rapid Exploration of Aging and Diseases in a Naturally Short-Lived Vertebrate

Abstract

Summary Aging is a complex process that affects multiple organs. Modeling aging and age-related diseases in the lab is challenging because classical vertebrate models have relatively long lifespans. Here we develop the first platform for rapid exploration of age-dependent traits and diseases in vertebrates, using the naturally short-lived African turquoise killifish. We provide an integrative genomic and genome-editing toolkit in this organism using our de novo-assembled genome and the CRISPR/Cas9 technology. We mutate many genes encompassing the hallmarks of aging, and for a subset, we produce stable lines within 2–3 months. As a proof-of-principle, we show that fish deficient for the protein subunit of telomerase exhibit the fastest onset of telomere-related pathologies among vertebrates. We further demonstrate the feasibility of creating specific genetic variants. This genome-to-phenotype platform represents a unique resource for studying vertebrate aging and disease in a high throughput manner and for investigating candidates arising from human genome-wide studies.

The African turquoise killifish: A research organism to study vertebrate aging and diapause

“…Transposase Tol2‐based transgenesis was successfully developed in turquoise killifish (Allard, Kamei & Duan, 2013; Hartmann & Englert, 2012; Valenzano et al., 2011). Importantly, the Brunet laboratory at Stanford recently developed a highly efficient CRISPR‐Cas9 genome‐editing pipeline, which allows both knockout and knockin in this fish with very high efficiency (Harel et al., 2015; Figure 3). Given the short lifecycle of the turquoise killifish, a stable line with the desired modification can be generated in 2 to 3 months (Harel et al., 2015), which is faster and more advantageous than both zebrafish and mice especially when the time for back‐crossing is also considered.…”

“…For example, while a 3‐month‐old turquoise killifish of the GRZ strain is entering old age (Table 1), a 3‐month‐old zebrafish just reaches sexual maturation to breed (Avdesh et al., 2012). The turquoise killifish thus provides a fast and versatile genotype‐to‐phenotype platform, which is especially helpful for studying aging‐related diseases, including cancer and neurodegeneration (Di Cicco et al., 2011; Harel et al., 2015; Terzibasi Tozzini et al., 2014; Tozzini et al., 2012). A good example is the telomerase reverse transcriptase ( TERT ), which when mutated in the turquoise killifish, results in phenotypes that are similar to those seen in humans deficient for telomerase: defects in actively proliferating organs and systems, such as blood, gut, and gonads (Harel et al., 2015; Figure 3).…”

“…The turquoise killifish thus provides a fast and versatile genotype‐to‐phenotype platform, which is especially helpful for studying aging‐related diseases, including cancer and neurodegeneration (Di Cicco et al., 2011; Harel et al., 2015; Terzibasi Tozzini et al., 2014; Tozzini et al., 2012). A good example is the telomerase reverse transcriptase ( TERT ), which when mutated in the turquoise killifish, results in phenotypes that are similar to those seen in humans deficient for telomerase: defects in actively proliferating organs and systems, such as blood, gut, and gonads (Harel et al., 2015; Figure 3). Importantly, the telomeres in the turquoise killifish are more comparable in length to human telomeres (~8 kb) than mouse telomeres (which are extremely long ~50–150 kb; Harel et al., 2015; Hartmann et al., 2009), making this fish a great research system to model and study telomere attrition in the context of aging and telomere‐related pathologies (e.g., age‐dependent stem cell exhaustion) in vivo.…”

High telomerase is a hallmark of undifferentiated spermatogonia and is required for maintenance of male germline stem cells

“…Telomere shortening in mice or fish lacking telomerase causes infertility and the eventual loss of male germ cells (Lee et al 1998;Henriques et al 2013;Harel et al 2015). Inactivation of telomerase in worms leads to infertility and a loss of male germ cells as telomeres shorten (Meier et al 2006).…”

Short-lived fish may hold clues to human ageing

“…These tests are already under way, says Anne Brunet, a geneticist at Stanford University in California and an author on one of the studies. Earlier this year, Brunet's team used CRISPR-Cas9 genome editing in 'proof-of-principle' experiments to alter several ageing-related genes in killifish 3 . "We are excited at trying to make it live longer, " she says.…”