Concerted evolution reveals co-adapted amino acid substitutions in Na+K+-ATPase of frogs that prey on toxic toads

Mohammadi, Shabnam; Yang, Lu; Harpak, Arbel; Herrera‐Álvarez, Santiago; Rodríguez-Ordóñez, María del Pilar; Peng, Julie; Zhang, Karen; Storz, Jay F.; Dobler, Susanne; Crawford, Andrew J.; Andolfatto, Peter

doi:10.1016/j.cub.2021.03.089

Cited by 28 publications

(57 citation statements)

References 67 publications

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“…Because ion transport across the membrane is a primary function of NKA and its disruption can have severe pathological effects [36], mutations that compromise this function are likely to be under strong purifying selection. As suggested by previous work [23][24][25], CTS-resistant substitutions at sites 111 and 122 can decrease enzyme activity. We evaluated the generality of these effects across a broader phylogenetic scale by comparing enzyme activity of the 15 mutant NKA proteins to their corresponding wild-type proteins.…”

Section: Pleiotropic Effects On Nka Activity Largely Depend On States...supporting

confidence: 65%

“…2). Most of the variation in ATP1A2 at these sites is restricted to squamate reptiles and ATP1A3 lacks substitutions at site 122 altogether, despite the well-known potential for substitutions at this site to confer CTS resistance [25,31]. Looking across species and paralogs, the extent of convergence at sites 111 and 122 is remarkable (Figs.…”

Section: Patterns Of Atp1a Sequence Evolution Across Species and Para...mentioning

confidence: 99%

“…Functional investigations of CTS resistance-conferring substitutions at two key sites (111 and 122) in ATP1A orthologs of Drosophila [23,24] and Neotropical grass frogs [25] revealed associated negative pleiotropic effects on protein function and showed that evolution at other sites in the protein mitigate these detrimental effects. In light of these pleiotropic and epistatic constraints, it is curious that convergent CTS-resistant substitutions are often observed among ATP1A orthologs of highly divergent species.…”

Section: Introductionmentioning

confidence: 99%

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Constraints on the evolution of toxin-resistant Na,K-ATPases have limited dependence on sequence divergence

Mohammadi

Yang

Herrera‐Álvarez

et al. 2021

Preprint

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Comparative genomic studies reveal a global decline in rates of convergent amino acid substitution as a function of evolutionary distance. This pattern has been attributed to epistatic constraints on protein evolution, the idea being that mutations tend to confer the same fitness effects on more similar genetic backgrounds, so convergent substitutions are more likely to occur in closely related species. However, this hypothesis lacks experimental validation. We tested this model in the context of the recurrent evolution of resistance to cardiotonic steroids (CTS) across diverse groups of tetrapods, which occurs via specific amino acid substitutions to the α-subunit family of Na+,K+-ATPases (ATP1A). After identifying a series of recurrent substitutions at two key sites of ATP1A1 predicted to confer CTS resistance, we performed protein engineering experiments to test the functional consequences of introducing these substitutions onto divergent species backgrounds. While we find that substitutions at these sites can have substantial background-dependent effects on CTS resistance, we also find no evidence for background-dependent effects on protein activity. We further show that the magnitude of a substitution’s effect on activity does not depend on the overall extent of ATP1A1 sequence divergence between species. More generally, a global analysis of substitution patterns across ATP1A orthologs and paralogs reveals that the probability of convergent substitution protein-wide is not predicted by sequence divergence. Together, these findings suggest that intramolecular epistasis is not an important constraint on the evolution of ATP1A CTS resistance in tetrapods.Significance StatementIndividual amino acid residues within a protein work in concert to produce a functionally coherent structure that must be maintained even as orthologous proteins in different species diverge over time. Given this dependence, we expect identical mutations to have more similar effects on protein function in more closely related species. We tested this hypothesis by performing protein-engineering experiments on ATP1A, an enzyme mediating target-site insensitivity to cardiotonic steroids (CTS) in diverse animals. These experiments reveal that although the phenotypic effects of substitutions can sometimes be background-dependent, the magnitude of these effects does not correlate with ATP1A1 sequence divergence. This implies that the genetic background across the ATP1A protein does not strongly limit the evolution of CTS resistance in animals.

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Section: Pleiotropic Effects On Nka Activity Largely Depend On States...supporting

confidence: 65%

Section: Patterns Of Atp1a Sequence Evolution Across Species and Para...mentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Constraints on the evolution of toxin-resistant Na,K-ATPases have limited dependence on sequence divergence

Mohammadi

Yang

Herrera‐Álvarez

et al. 2021

Preprint

Self Cite

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“…Thus, a possible evolutionary pathway for bufadienolide defense in toads is via natural selection on the regulation of endogenous CGs ( Flier et al, 1980 ) coupled with the development of Na + /K + -ATPase target site insensitivity, whereby amino acid substitutions result in a weaker affinity of Na + /K + -ATPase for CGs. Target site insensitivity to CGs has been demonstrated in the α3 Na + /K + -ATPase subunit of bufonid toads – including Atelopus spumarius – and toad-feeding reptiles ( Moore et al, 2009 ; Ujvari et al, 2015 ) and in a tandem duplicate of the α1 Na + /K + -ATPase in toad-feeding frogs ( Leptodactylus; Mohammadi et al, 2021 ). More than one hundred different bufadienolides have been detected in the skins, eggs, or granular gland secretions of bufonid toads ( Rodríguez et al, 2017 ).…”

Section: Atelopus Toxins – Chemical Structures Pharmacology ...mentioning

confidence: 99%

A review of chemical defense in harlequin toads (Bufonidae: Atelopus)

Pearson

Tarvin

2022

Toxicon: X

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“…Zwei davon, Q111R und N122D, befinden sich in der extrazellulären Schleife der αUntereinheit von ATP1A1, an die die Bufadienolide binden. Anhand von gereinigten en zymvarianten, die die verschiedenen Austausche in unterschiedlichen Kom binationen trugen, konnten amerika nische Wissenschaftler die Bedeutung von Q111R und N122D für die Toxin resistenz zeigen [2]. Aber welche Rolle mit ihrer behäbigen Fortbewegungs weise erscheinen Kröten wie die harmlosigkeit schlechthin.…”

Section: Resistenz Ohne Nebenwirkungenunclassified

Die stumpfe Waffe der Kröten

Tetsch¹

2021

Chemie in nserer Zeit

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Concerted evolution reveals co-adapted amino acid substitutions in Na+K+-ATPase of frogs that prey on toxic toads

Cited by 28 publications

References 67 publications

Constraints on the evolution of toxin-resistant Na,K-ATPases have limited dependence on sequence divergence

Constraints on the evolution of toxin-resistant Na,K-ATPases have limited dependence on sequence divergence

A review of chemical defense in harlequin toads (Bufonidae: Atelopus)

Die stumpfe Waffe der Kröten

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