Evolution of diapause in the African turquoise killifish by remodeling ancient gene regulatory landscape

Abstract

Suspended animation states such as hibernation or diapause allow organisms to survive extreme environments. But the mechanisms underlying the evolution of these extreme survival states are unknown. The African turquoise killifish has evolved diapause as a form of suspended development to survive the complete drought that occurs every year in its habitat. Here we show that many gene duplicates - paralogs - exhibit specialized expression in diapause versus normal development in the African turquoise killifish. Surprisingly, paralogs with specialized expression in diapause are evolutionarily very ancient, and they are also present even in vertebrates that do not exhibit diapause. Profiling the chromatin accessibility landscape among different fish species reveals an evolutionarily recent increase in chromatin accessibility at these very ancient paralogs, suggesting rewiring of their regulatory landscape. The increase in chromatin accessibility in the African turquoise killifish is linked to the presence of new binding sites for transcription factors (e.g., FOXO, REST, and PPAR), due to both de novo mutations and transposable element insertion. Interestingly, accessible chromatin regions in diapause are enriched for lipid metabolism genes. By performing lipidomics in different fish species, we uncover a specific lipid profile in African turquoise killifish embryos in diapause. Notably, select very long-chain fatty acids are high in diapause, suggesting they may be used for long-term survival in this state. Together, our multi-omic analysis indicates that diapause is driven by regulatory innovation of very ancient gene programs that are critical for survival. Our work also suggests a mechanism for how complex adaptations evolve in nature and offers strategies by which a suspended animation program could be reactivated in other species for long-term preservation.

Metabolic enhancement of mammalian developmental pausing

“…Forkhead box (FOX) family transcription factors (known as DAF16 in C.elegans) regulate cellular metabolism, nutrient stress, longevity and lifespan in multiple species 29-31 and show dynamic cellular localization in response to cellular signaling 32,33 . In addition, FOXO proteins have been proposed to play a role in marsupial and killifish diapause 34,35 .…”

“…Lipid reserves and FAO appear to play an essential role in regulating the balance of stem cell dormancy and proliferation in several adult tissues 8,[43][44][45][46] , as well as during embryonic diapause across species 35,[47][48][49] , although a contrary observation has been made in human cells 26 . Interestingly, enhanced lipid consumption due to dietary restriction increases life span in Drosophila 50 and C.elegans 51 .…”

“…In addition, FOXO proteins ! 10 have been proposed to play a role in marsupial and killifish diapause 34,35 .…”

Rapid and precise genome engineering in a naturally short-lived vertebrate

“…Tools to advance genetic interrogation of the killifish have been developed, including a sequenced genome (Reichwald et al, 2015; Valenzano et al, 2015) and Tol2 transgenesis (Allard et al, 2013; Hartmann and Englert, 2012; Valenzano et al, 2011), as well as CRISPR/Cas9-mediated knock-out (Harel et al, 2015) and CRISPR/Cas13-mediated knock-down (Kushawah et al, 2020). This genetic toolkit has enabled discoveries about the mechanisms of aging (Astre et al, 2022a; Bradshaw et al, 2022; Chen et al, 2022; Harel et al, 2022; Louka et al, 2022; Matsui et al, 2019; Smith et al, 2017; Van Houcke et al, 2021b), regeneration (Vanhunsel et al, 2022a; Vanhunsel et al, 2021; Vanhunsel et al, 2022b; Wang et al, 2020), evolution (Cui et al, 2019; Sahm et al, 2017; Singh et al, 2021; Willemsen et al, 2020), development (Abitua et al, 2021; Dolfi et al, 2019), and ‘suspended animation’ (Hu et al, 2020; Singh et al, 2021).…”

Rapid and precise genome engineering in a naturally short-lived vertebrate

“…Tools to advance genetic interrogation of the killifish have been developed, including a sequenced genome ( Reichwald et al, 2015 ; Valenzano et al, 2015 ), Tol2 transgenesis ( Allard et al, 2013 ; Hartmann and Englert, 2012 ; Valenzano et al, 2011 ), CRISPR/Cas9-mediated knock-out ( Harel et al, 2015 ), and CRISPR/Cas13-mediated knock-down ( Kushawah et al, 2020 ). This genetic toolkit has enabled discoveries about the mechanisms of aging ( Astre et al, 2022a ; Bradshaw et al, 2022 ; Chen et al, 2022 ; Harel et al, 2022 ; Louka et al, 2022 ; Matsui et al, 2019 ; Smith et al, 2017 ; Van Houcke et al, 2021b ; Vanhunsel et al, 2021 ), regeneration ( Vanhunsel et al, 2022a ; Vanhunsel et al, 2022b ; Wang et al, 2020 ), evolution ( Cui et al, 2019 ; Sahm et al, 2017 ; Singh et al, 2021 ; Willemsen et al, 2020 ), development ( Abitua et al, 2021 ; Dolfi et al, 2019 ), and embryonic diapause – a state of ‘suspended animation’ ( Hu et al, 2020 ; Singh et al, 2021 ).…”