Event Abstract Back to Event The molecular basis of pelvic brooding, a novel reproductive strategy Leon Hilgers1*, Arne W. Nolte2, Tobias Spanke1, Jana M. Flury1, Fabian Herder1, Bernhard Misof1 and Julia Schwarzer1 1 Zoological Research Museum Alexander Koenig (LG), Germany 2 University of Oldenburg, Germany Although evolutionary innovations have fascinated evolutionary biologists for more than 150 years, our understanding of their molecular underpinnings is still in its infancy. Novel reproductive strategies represent particularly promising models to study evolutionary innovation, because they rely on complex sets of behavioral, physiological and anatomical adaptations and sometimes even involve the origin of novel organs. Sulawesi ricefishes (Actinopterygii: Adrianichthyidae) exhibit a novel reproductive strategy that is referred to as pelvic brooding 1–3. Females of pelvic brooding species carry their clutch in a ventral concavity, covered by their pelvic fins, until the fry hatches. The eggs are connected to the female by attaching filaments, which are anchored inside the abdomen by a unique structure, called the plug 4. Furthermore, ovulation is delayed in pelvic brooders as long as eggs are attached and accelerated in case eggs are prematurely lost 5. Thus, pelvic brooding likely relies on several adaptations such as modified fins, shortened ribs, altered timing of ovulation and a novel plug structure 2–5. Despite its apparent complexity, pelvic brooding likely evolved from oviparous ancestors in at least two lineages 6. While the potentially repeated and likely recent origin of pelvic brooding makes it a promising system to study evolutionary innovations, its molecular basis remained so far unexplored. Here we use tissue-specific transcriptomes of brooding and non-brooding Oryzias eversi to gain insight into the genetic basis of pelvic brooding. We combine differential expression and functional enrichment analyses to shed light on pelvic-brooding related gene regulation and dominant molecular functions in different tissues including the novel plug structure. To this end, brooding females of O. eversi were separated and tissues harvested during mid-brooding and after brooding was completed when fish can, in principle, spawn again. Here we show that egg-carrying is associated with significant tissue-specific changes in gene expression. Changes in gene expression are strongest in the ovary with 47 differentially expressed (DE) genes (false discovery rate (FDR) ≤ 0.001), followed by the papilla (18 DE genes) and least severe in the skin (3 DE genes), which served as a reference tissue. Gene expression in the ovary is mostly down-regulated during brooding, matching suppressed ovulation during egg-carrying. Although gene expression between the dermally derived structures papilla and skin is very similar, it becomes more distinct during brooding. These findings in combination with significant gene expression changes in the papilla corroborate the morphology-based hypothesis that the genital papilla may play a role in pelvic brooding. Furthermore, the plug, which is a novel tissue that forms after spawning, exhibits remarkably distinct gene expression compared to the other tissues. Genes with plug-specific expression (overexpressed against all other tissues, FDR ≤ 10-10) are significantly enriched in genes with functions in immunity and include genes that also have important functions in mammalian placentas. Hence, our results indicate an important role for the immune system in plug formation during pelvic brooding and substantiate some parallels of pelvic-brooding with mammalian pregnancy and placentation. Future studies will build upon this work to investigate to what extend tissue-specific changes in gene expression during brooding are based on regulatory evolution, whether or not pelvic brooding has evolved in parallel and if so, to what extend similar molecular mechanisms underly its repeated evolution. In conclusion, this study sheds first light on the molecular basis of pelvic brooding and paves the way for a better understanding of the genetic basis of novel reproductive strategies and ultimately evolutionary innovations.
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