Expression profiling by high throughput sequencing
Summary
Motherhood is characterized by dramatic changes in brain and behavior, but less is known about fatherhood. Here we report that male sticklebacks – a small fish in which fathers provide care – experience dramatic changes in neurogenomic state as they become fathers. Some genes are unique to different stages of paternal care, some genes are shared across stages, and some genes are added to the previously acquired neurogenomic state. Comparative genomic analysis suggests that some of these neurogenomic dynamics resemble changes associated with pregnancy and reproduction in mammalian mothers. Moreover, gene regulatory analysis identified transcription factors that are regulated in opposite directions in response to a territorial challenge versus during paternal care. Altogether these results show that some of the molecular mechanisms of parental care might be deeply conserved and might not be sex-specific, and suggest that tradeoffs between opposing social behaviors are managed at the gene regulatory level.
Overall design
In order to track transcriptional dynamics associated with becoming a father, we sampled males for brain gene expression profiling at five different points during the reproductive cycle (n=5 males per time point): nest, eggs, early hatching, middle hatching and late hatching (control: reproductively mature males with no nests). Males in the nest condition had a nest but had not yet mated. Males in the eggs condition were sampled four days after their eggs were fertilized. Because males in the eggs condition were sampled four days after mating, the transcriptomic effects of mating are likely to have attenuated by the time males were sampled at this stage. Hatching takes place over the course of the fifth day after fertilization, and a previous study found that brain activation as assessed by Egr-1 expression was highest while male sticklebacks were caring for fry as compared to males with nests or eggs. In order to capture males’ response to the new social stimulus of their fry, we focused on three time points on the day of hatching which capture the start of the hatching process (9am), when approximately half of the clutch is hatched (1pm) and when all of the eggs have hatched (5pm). Males in the nest, eggs and early hatching conditions were sampled at 9am, males in the mid-hatching condition were sampled at 1pm and males in the late hatching condition were sampled at 5pm. Males in these conditions were compared to reproductively mature circadian-matched control males that did not have a nest (n=5 males per control group). Wild-caught females from the same population were used as mothers. Males were quickly netted and sacrificed by decapitation within seconds. All methods were approved by the IACUC of the University of Illinois at Urbana-Champaign (#15077).
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