Generation of a transparent killifish line through multiplex CRISPR/Cas9-mediated gene inactivation

Abstract

Body pigmentation is a major limitation for in vivo imaging and thus for the performance of longitudinal studies in biomedicine. A possibility to circumvent this obstacle is the employment of pigmentation mutants, which are used in fish species like zebrafish and medaka. To address the molecular basis of aging, the short-lived African killifish Nothobranchius furzeri has recently been established as a model organism. Despite its short lifespan, N. furzeri shows typical signs of mammalian aging including telomere shortening, accumulation of senescent cells and loss of regenerative capacity. Here, we report the generation of a transparent N. furzeri line by simultaneous inactivation of three key loci responsible for pigmentation. We demonstrate that this stable line, named klara, can serve as a tool for different in vivo applications including behavioral experiments addressing mate choice and the establishment of a senescence reporter by homology-directed repair-mediated integration of a fluorophore into the cdkn1a (p21) locus.

A Year at the Forefront of Proteostasis and Aging

“…Advances in knock-in technology using CRISPR were recently obtained in the African turquoise killifish, a vertebrate species that has a short lifespan of just 4-6 months ( Nath et al, 2022 preprint). By mutating genes involved in body pigmentation, a transparent killifish reporter line to monitor senescence was created, which could be used to identify new anti-aging interventions ( Krug et al, 2022 preprint). Extremophile organisms are characterised by their high resistance to environmental stress, and their study could bring novel insights into the maintenance of proteostasis.…”