The transcriptomic evolution of mammalian pregnancy:gene expression innovations in endometrial stromal fibroblasts

The endometrial stromal fibroblast (ESF) is a cell type present in the uterine lining of therian mammals. In the stem lineage of eutherian mammals, ESF acquired the ability to differentiate into decidual cells in order to allow embryo implantation. We call the latter cell type “neo-ESF” in contrast to “paleo-ESF” which is homologous to eutherian ESF but is not able to decidualize. In this study, we compare the transcriptomes of ESF from six therian species: Opossum (Monodelphis domestica; paleo-ESF), mink, rat, rabbit, human (all neo-ESF), and cow (secondarily nondecidualizing neo-ESF). We find evidence for strong stabilizing selection on transcriptome composition suggesting that the expression of approximately 5,600 genes is maintained by natural selection. The evolution of neo-ESF from paleo-ESF involved the following gene expression changes: Loss of expression of genes related to inflammation and immune response, lower expression of genes opposing tissue invasion, increased markers for proliferation as well as the recruitment of FOXM1, a key gene transiently expressed during decidualization. Signaling pathways also evolve rapidly and continue to evolve within eutherian lineages. In the bovine lineage, where invasiveness and decidualization were secondarily lost, we see a re-expression of genes found in opossum, most prominently WISP2, and a loss of gene expression related to angiogenesis. The data from this and previous studies support a scenario, where the proinflammatory paleo-ESF was reprogrammed to express anti-inflammatory genes in response to the inflammatory stimulus coming from the implanting conceptus and thus paving the way for extended, trans-cyclic gestation

Transposable elements and host evolution: A case of transposon-mediated regulatory innovation during the evolution of pregnancy

Organisms evolve by changes to coding and regulatory DNA. To understand how organismal phenotypes arise and diverge over time, therefore, it is necessary to elucidate the mechanisms underlying coding and regulatory evolution. Studies on regulatory evolution have provided insights into how regulatory elements change or are lost over time, but how new elements—such as promoters and enhancers—originate is an open question. One hypothesis is that new regulatory elements are generated de novo by the random gain of transcription factor binding sites (TFBSs) near genes. An alternative opinion, first proposed decades ago, is that new regulatory elements are co-opted from transposable elements as "ready-to-use" elements. Transposable elements (TEs), mobile DNA elements that invade and replicate within genomes, comprise about half of mammalian genomes and contain a variety of regulatory signals necessary for their own propagation. Thus, TEs have the potential to impact host regulatory evolution, and ultimately phenotypic evolution, in a significant way. In this dissertation I investigate the role of transposable elements in the regulatory evolution of decidual prolactin (dPrl) in mammals, as the promoter in primates derives from a lineage-specific TE called MER39. To understand if and how MER39 has affected dPrl regulation and endometrial functioning in mammals and primates in particular, I surveyed its expression during pregnancy in various mammals and the location(s) of transcriptional initiation. I found that Prl was convergently recruited into uterine expression in primates, rodents, and elephants by the co-option of different transposable elements, highlighting the frequency at which TEs are used for regulatory functions by the host and their importance in regulatory innovation. I also traced the origin of the MER39-derived promoter in primates, showing that evolution of the strong dPrl promoter in apes was a multistep process that took millions of years. Strong promoter activity of MER39 evolved coincident with the origin of a novel reproductive character in apes, interstitial invasion; thus, transformation of this TE into a regulatory element likely played a role in the evolution of pregnancy in apes. Mechanistically, I show that strong promoter activity in apes involves epistatic interactions between TFBSs ancestral to MER39 and derived sites. I propose a novel mode of molecular evolution by which MER39 was transformed, called "epistatic capture," defined as the fixation of a TFBS that is ancestral but variable in outgroup lineages, and is fixed in the ingroup because of epistatic interactions with derived TFBSs. A review of the literature suggests that epistatic capture may be a common mechanism by which TEs are domesticated for regulatory functions in host tissues like the endometrium. Finally, since TEs have had a major impact on regulatory and other types of innovations in placental tissues, I argue that TEs have facilitated the rampant diversification of the placenta in eutherian mammals and potentially other fast-evolving tissues

Adaptive changes in the transcription factor HoxA-11 are essential for the evolution of pregnancy in mammals

Evolutionary change in gene regulation can result from changes in cis-regulatory elements, leading to differences in the temporal and spatial expression of genes or in the coding region of transcription factors leading to novel functions or both. Although there is a growing body of evidence supporting the importance of cis-regulatory evolution, examples of protein-mediated evolution of novel developmental pathways have not been demonstrated. Here, we investigate the evolution of prolactin (PRL) expression in endometrial cells, which is essential for placentation/pregnancy in eutherian mammals and is a direct regulatory target of the transcription factor HoxA-11. Here, we show that (i) endometrial PRL expression is a derived feature of placental mammals, (ii) the PRL regulatory gene HoxA-11 experienced a period of strong positive selection in the stem-lineage of eutherian mammals, and (iii) only HoxA-11 proteins from placental mammals, including the reconstructed ancestral eutherian gene, are able to up-regulate PRL from the promoter used in endometrial cells. In contrast, HoxA-11 from the reconstructed therian ancestor, opossum, platypus, and chicken are unable to up-regulate PRL expression. These results demonstrate that the evolution of novel gene expression domains is not only mediated by the evolution of cis-regulatory elements but can also require evolutionary changes of transcription factor proteins themselves

Massively parallel disruption of enhancers active in human neural stem cells

Summary: Changes in gene regulation have been linked to the expansion of the human cerebral cortex and to neurodevelopmental disorders, potentially by altering neural progenitor proliferation. However, the effects of genetic variation within regulatory elements on neural progenitors remain obscure. We use sgRNA-Cas9 screens in human neural stem cells (hNSCs) to disrupt 10,674 genes and 26,385 conserved regions in 2,227 enhancers active in the developing human cortex and determine effects on proliferation. Genes with proliferation phenotypes are associated with neurodevelopmental disorders and show biased expression in specific fetal human brain neural progenitor populations. Although enhancer disruptions overall have weaker effects than gene disruptions, we identify enhancer disruptions that severely alter hNSC self-renewal. Disruptions in human accelerated regions, implicated in human brain evolution, also alter proliferation. Integrating proliferation phenotypes with chromatin interactions reveals regulatory relationships between enhancers and their target genes contributing to neurogenesis and potentially to human cortical evolution