Genetic and environmental regulation of breeding and migratory behavior in wild birds: an experimental approach

Assessing how genes and environmental cues interact to shape phenotypic variation is a key issue of evolutionary biology. The same genotype may in fact produce different phenotypes when experiencing different environmental conditions, resulting in so-called reaction norms or gene-environment interactions, that represent major sources of phenotypic variation and are fundamental for adaptation and evolution. In migratory species, traits related to migration and breeding show large phenotypic and additive genetic variance. Recently, several phenological candidate genes (PCG), mostly related to the control of circadian and seasonal clocks, have been suggested to play an important role in shaping migration and breeding traits of vertebrates, including avian taxa. On the other hand, these traits also show a large environmental component. Such broad environmental variation in migration and breeding traits may be associated with epigenetic DNA modifications, such as DNA methylation, which is believed to be a major source of phenotypic plasticity in natural populations, shifting phenotypic traits depending on contingent and/or past environmental conditions. Moreover, the interactive effects of PCG and environmental conditions in shaping migration and breeding traits remain largely elusive, and have never been experimentally investigated. By exploiting the most advanced animal tracking technologies, such as satellite tracking and light-level geolocation, we plan to elucidate how the interaction between environmental conditions and PCG variability acts to shape behavioral traits of two long-distance, tropical wintering migratory bird species (the lesser kestrel, Falco naumanni and the Scopoli’s shearwater, Calonectris diomedea) through the entire annual cycle. We will first investigate the carry-over effects of an experimental alteration of the breeding environment on migration and wintering, next season breeding behavior and reproductive investment. We plan to characterize the individual genetic background at a suite of PCG likely to modulate breeding and migratory behavior (i.e. ADCYAP1, CREB1, CLOCK, NPAS2), and to estimate individual-level multilocus heterozygosity via high-throughput sequencing techniques, with the aim of investigating whether PCG variation and multilocus heterozygosity predict migration and breeding traits. Finally, we propose to analyze the extent to which DNA methylation patterns of PCG, avian glucocorticoid receptors and highly conserved transposable elements scattered throughout the avian genome (as markers of overall methylation at the genomic level) affect phenotypic traits expression and fitness, accounting for genetic background, and are modified by experimental modification of ecological conditions during breeding. To our knowledge, this project constitutes the first attempt to experimentally evaluate PCG-environment interactions in wild birds. We are confident to gain key novel insights into: 1) how manipulating the breeding environment carries over to subsequent major life-cycle events and fitness traits; 2) the extent to which PCG, multilocus heterozygosity, and methylation affect migration and breeding traits; 3) how PCG- and multilocus heterozygosity-environment interactions shape migration and breeding traits; 4) how DNA methylation (both at the genomic level and at specific genes) is affected by an altered breeding environment and whether it predicts migration and fitness traits. We anticipate that, by experimentally investigating the effects of PCG-environment interactions and genomic/gene-specific methylation levels on migratory and breeding behavior, the project is going to produce major advances in assessing the evolutionary potential of migratory traits in natural populations, hence greatly improving our understanding of how long-distance migratory birds face the challenges imposed by unpredictable environmental variation.