Research Article

A genetic signature of the evolution of loss of flight in the Galapagos cormorant

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Science  02 Jun 2017:
Vol. 356, Issue 6341, eaal3345
DOI: 10.1126/science.aal3345

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Loss of flight in the Galapagos cormorant

Although rare among existing birds, the loss of flight appears to have occurred multiple times in evolutionary history. However, the genetic changes that ground avian species are not well understood. Burga et al. sequenced genomes from three cormorant species and compared them with that of the flightless Galapagos cormorant (see the Perspective by Cooper). They identified variants in genes involved in primary ciliogenesis. Functional analyses of these variants suggest that the impaired function of the genes may be responsible for skeletal changes associated with the Galapagos cormorant’s loss of flight.

Science, this issue p. eaal3345; see also p. 904

Structured Abstract

INTRODUCTION

Changes in the size and proportion of limbs and other structures have played a key role in the evolution of species. One common class of limb modification is recurrent wing reduction and loss of flight in birds. Indeed, Darwin used the occurrence of flightless birds as an argument in favor of his theory of natural selection. Loss of flight has evolved repeatedly and is found among 26 families of birds in 17 different orders. Despite the frequency of these modifications, we have a limited understanding of their underpinnings at the genetic and molecular levels.

RATIONALE

To better understand the evolution of changes in limb size, we studied a classic case of recent loss of flight in the Galapagos cormorant (Phalacrocorax harrisi). Cormorants are large water birds that live in coastal areas or near lakes, and P. harrisi is the only flightless cormorant among approximately 40 extant species. The entire population is distributed along the coastlines of Isabela and Fernandina islands in the Galapagos archipelago. P. harrisi has a pair of short wings, which are smaller than those of any other cormorant. The extreme reduction of the wings and pectoral skeleton observed in P. harrisi is an attractive model for studying the evolution of loss of flight because it occurred very recently; phylogenetic evidence suggests that P. harrisi diverged from its flighted relatives within the past 2 million years. We developed a comparative and predictive genomics approach that uses the genome sequences of P. harrisi and its flighted relatives to find candidate genetic variants that likely contributed to the evolution of loss of flight.

RESULTS

We sequenced and de novo assembled the whole genomes of P. harrisi and three closely related flighted cormorant species. We identified thousands of coding variants exclusive to P. harrisi and classified them according to their probability of altering protein function based on conservation. Variants most likely to alter protein function were significantly enriched in genes mutated in human skeletal ciliopathies, including Ofd1, Evc, Wdr34, and Ift122. We carried out experiments in Caenorhabditis elegans to confirm that a missense variant present in the Galapagos cormorant IFT122 protein is sufficient to affect ciliary function. The primary cilium is essential for Hedgehog (Hh) signaling in vertebrates, and individuals affected by ciliopathies have small limbs and ribcages, mirroring the phenotype of P. harrisi. We also identified a 4–amino acid deletion in the regulatory domain of Cux1, a highly conserved transcription factor that has been experimentally shown to regulate limb growth in chicken. The four missing amino acids are perfectly conserved in all birds and mammals sequenced to date. We tested the consequences of this deletion in a chondrogenic cell line and showed that it impairs the ability of CUX1 to transcriptionally up-regulate cilia-related genes (some of which contain function-altering variants in P. harrisi) and to promote chondrogenic differentiation. Finally, we show that positive selection may have played a role in the fixation of the variants associated with loss of flight in P. harrisi.

CONCLUSION

Our results indicate that the combined effect of variants in genes necessary for the correct transcriptional regulation and function of the primary cilium likely contributed to the evolution of highly reduced wings and other skeletal adaptations associated with loss of flight in P. harrisi. Our approach may be generally useful for identification of variants underlying evolutionary novelty from genomes of closely related species.

Comparative and predictive genomics of loss of flight.

Comparison of the genomes of four closely related cormorant species allowed us to predict function-altering variants exclusively affecting the Galapagos cormorant and to test their functional consequences. Our results implicate ciliary dysfunction as a likely contributor to the evolution of loss of flight.

Abstract

We have a limited understanding of the genetic and molecular basis of evolutionary changes in the size and proportion of limbs. We studied wing and pectoral skeleton reduction leading to flightlessness in the Galapagos cormorant (Phalacrocorax harrisi). We sequenced and de novo assembled the genomes of four cormorant species and applied a predictive and comparative genomics approach to find candidate variants that may have contributed to the evolution of flightlessness. These analyses and cross-species experiments in Caenorhabditis elegans and in chondrogenic cell lines implicated variants in genes necessary for transcriptional regulation and function of the primary cilium. Cilia are essential for Hedgehog signaling, and humans affected by skeletal ciliopathies suffer from premature bone growth arrest, mirroring skeletal features associated with loss of flight.

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