Deena Emera

The Evolution of Lineage-Specific Regulatory Activities in the Human Embryonic Limb

The evolution of human anatomical features likely involved changes in gene regulation during development. However, the nature and extent of human-specific developmental regulatory functions remain unknown. We obtained a genome-wide view of cis-regulatory evolution in human embryonic tissues by comparing the histone modification H3K27ac, which provides a quantitative readout of promoter and enhancer activity, during human, rhesus, and mouse limb development. Based on increased H3K27ac, we find that 13% of promoters and 11% of enhancers have gained activity on the human lineage since the human-rhesus divergence. These gains largely arose by modification of ancestral regulatory activities in the limb or potential co-option from other tissues and are likely to have heterogeneous genetic causes. Most enhancers that exhibit gain of activity in humans originated in mammals. Gains at promoters and enhancers in the human limb are associated with increased gene expression, suggesting they include molecular drivers of human morphological evolution.

Ancient transposable elements transformed the uterine regulatory landscape and transcriptome during the evolution of mammalian pregnancy

SUMMARY A major challenge in biology is determining how evolutionarily novel characters originate; however, mechanistic explanations for the origin of new characters are almost completely unknown. The evolution of pregnancy is an excellent system in which to study the origin of novelties because mammals preserve stages in the transition from egg laying to live birth. To determine the molecular bases of this transition, we characterized the pregnant/gravid uterine transcriptome from tetrapods to trace the evolutionary history of uterine gene expression. We show that thousands of genes evolved endometrial expression during the origins of mammalian pregnancy, including genes that mediate maternal-fetal communication and immunotolerance. Furthermore, thousands of cis-regulatory elements that mediate decidualization and cell-type identity in decidualized stromal cells are derived from ancient mammalian transposable elements (TEs). Our results indicate that one of the defining mammalian novelties evolved from DNA sequences derived from ancient mammalian TEs coopted into hormone-responsive regulatory elements distributed throughout the genome.

Evolutionary changes in promoter and enhancer activity during human corticogenesis

Of mice, men, and macaque brains The human brain represents a unique evolutionary trajectory. To better understand how the human brain came to be, Reilly et al. sought to identify changes in gene expression between mice, macaques, and humans. They compared epigenetic marks in the embryonic cortex, which revealed changes in gene regulation in biological pathways associated with cortical development. Science, this issue p. 1155 Epigenetic marks associated with evolution of the human cortex gained activity relative to those in mice and monkeys. Human higher cognition is attributed to the evolutionary expansion and elaboration of the human cerebral cortex. However, the genetic mechanisms contributing to these developmental changes are poorly understood. We used comparative epigenetic profiling of human, rhesus macaque, and mouse corticogenesis to identify promoters and enhancers that have gained activity in humans. These gains are significantly enriched in modules of coexpressed genes in the cortex that function in neuronal proliferation, migration, and cortical-map organization. Gain-enriched modules also showed correlated gene expression patterns and similar transcription factor binding site enrichments in promoters and enhancers, suggesting that they are connected by common regulatory mechanisms. Our results reveal coordinated patterns of potential regulatory changes associated with conserved developmental processes during corticogenesis, providing insight into human cortical evolution.

The evolution of menstruation: A new model for genetic assimilation

Why do humans menstruate while most mammals do not? Here, we present our answer to this long-debated question, arguing that (i) menstruation occurs as a mechanistic consequence of hormone-induced differentiation of the endometrium (referred to as spontaneous decidualization, or SD); (ii) SD evolved because of maternal-fetal conflict; and (iii) SD evolved by genetic assimilation of the decidualization reaction, which is induced by the fetus in non-menstruating species. The idea that menstruation occurs as a consequence of SD has been proposed in the past, but here we present a novel hypothesis on how SD evolved. We argue that decidualization became genetically stabilized in menstruating lineages, allowing females to prepare for pregnancy without any signal from the fetus. We present three models for the evolution of SD by genetic assimilation, based on recent advances in our understanding of the mechanisms of endometrial differentiation and implantation. Testing these models will ultimately shed light on the evolutionary significance of menstruation, as well as on the etiology of human reproductive disorders like endometriosis and recurrent pregnancy loss.

Beta-keratins of turtle shell are glycine-proline-tyrosine rich proteins similar to those of crocodilians and birds

This study presents, for the first time, sequences of five beta‐keratin cDNAs from turtle epidermis obtained by means of 5′‐ and 3′‐rapid amplification of cDNA ends (RACE) analyses. The deduced amino acid sequences correspond to distinct glycine‐proline‐serine‐tyrosine rich proteins containing 122–174 amino acids. In situ hybridization shows that beta‐keratin mRNAs are expressed in cells of the differentiating beta‐layers of the shell scutes. Southern blotting analysis reveals that turtle beta‐keratins belong to a well‐conserved multigene family. This result was confirmed by the amplification and sequencing of 13 genomic fragments corresponding to beta‐keratin genes. Like snake, crocodile and avian beta‐keratin genes, turtle beta‐keratins contain an intron that interrupts the 5′‐untranslated region. The length of the intron is variable, ranging from 0.35 to 1.00 kb. One of the sequences obtained from genomic amplifications corresponds to one of the five sequences obtained from cDNA cloning; thus, sequences of a total of 17 turtle beta‐keratins were determined in the present study. The predicted molecular weight of the 17 different deduced proteins range from 11.9 to 17.0 kDa with a predicted isoelectric point of 6.8–8.4; therefore, they are neutral to basic proteins. A central region rich in proline and with beta‐strand conformation shows high conservation with other reptilian and avian beta‐keratins, and it is likely involved in their polymerization. Glycine repeat regions, often containing tyrosine, are localized toward the C‐terminus. Phylogenetic analysis shows that turtle beta‐keratins are more similar to crocodilian and avian beta‐keratins than to those of lizards and snakes.

Transformation of a transposon into a derived prolactin promoter with function during human pregnancy

Transposable elements (TEs) are known to provide DNA for host regulatory functions, but the mechanisms underlying the transformation of TEs into cis-regulatory elements are unclear. In humans two TEs—MER20 and MER39—contribute the enhancer/promoter for decidual prolactin (dPRL), which is dramatically induced during pregnancy. We show that evolution of the strong human dPRL promoter was a multistep process that took millions of years. First, MER39 inserted near MER20 in the primate/rodent ancestor, and then there were two phases of activity enhancement in primates. Through the mapping of causal nucleotide substitutions, we demonstrate that strong promoter activity in apes involves epistasis between transcription factor binding sites (TFBSs) ancestral to MER39 and derived sites. We propose a mode of molecular evolution that describes the process by which MER20/MER39 was transformed into a strong promoter, called “epistatic capture.” Epistatic capture is the stabilization of a TFBS that is ancestral but variable in outgroup lineages, and is fixed in the ingroup because of epistatic interactions with derived TFBSs. Finally, we note that evolution of human promoter activity coincides with the emergence of a unique reproductive character in apes, highly invasive placentation. Because prolactin communicates with immune cells during pregnancy, which regulate fetal invasion into maternal tissues, we speculate that ape dPRL promoter activity evolved in response to increased invasiveness of ape fetal tissue.

Origin and evolution of developmental enhancers in the mammalian neocortex.

Significance The neocortex mediates complex cognitive and motor tasks in all mammals. A long-debated question is how this complex structure evolved in primitive mammals. Here we investigate the role of novel mammalian gene regulatory sequences in the emergence of the neocortex and the mechanisms by which these sequences emerged. We find that ∼20% of elements active during human and mouse neocortical development were born in early mammals. These novel mammalian elements enrich for cell migration, cell signaling, and axon guidance functions, implicating these processes in neocortical origins. In contrast to recent studies, we propose a model in which novel regulatory elements emerge as short sequences of minimal biological significance. Many disappear, but those that survive become increasingly complex over time. Morphological innovations such as the mammalian neocortex may involve the evolution of novel regulatory sequences. However, de novo birth of regulatory elements active during morphogenesis has not been extensively studied in mammals. Here, we use H3K27ac-defined regulatory elements active during human and mouse corticogenesis to identify enhancers that were likely active in the ancient mammalian forebrain. We infer the phylogenetic origins of these enhancers and find that ∼20% arose in the mammalian stem lineage, coincident with the emergence of the neocortex. Implementing a permutation strategy that controls for the nonrandom variation in the ages of background genomic sequences, we find that mammal-specific enhancers are overrepresented near genes involved in cell migration, cell signaling, and axon guidance. Mammal-specific enhancers are also overrepresented in modules of coexpressed genes in the cortex that are associated with these pathways, notably ephrin and semaphorin signaling. Our results also provide insight into the mechanisms of regulatory innovation in mammals. We find that most neocortical enhancers did not originate by en bloc exaptation of transposons. Young neocortical enhancers exhibit smaller H3K27ac footprints and weaker evolutionary constraint in eutherian mammals than older neocortical enhancers. Based on these observations, we present a model of the enhancer life cycle in which neocortical enhancers initially emerge from genomic background as short, weakly constrained “proto-enhancers.” Many proto-enhancers are likely lost, but some may serve as nucleation points for complex enhancers to evolve.

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.

An evolutionary test of the isoform switching hypothesis of functional progesterone withdrawal for parturition: humans have a weaker repressive effect of PR-A than mice

Abstract Background: A decrease in maternal serum progesterone (P4) concentrations precedes the onset of labor in most placental mammals. Humans differ by maintaining high levels of P4 throughout birth. Parturition in humans probably includes mechanisms that undercut the pregnancy sustaining function of P4. One attractive hypothesis is the isoform switching hypothesis (ISH). ISH is supported by in vitro evidence that progesterone receptor isoform A (PR-A) inhibits PR-B and that the PR-A/PR-B ratio increases towards term. Materials and methods: Here, we test the hypothesis that isoform switching is an adaptation to high levels of P4 at term, predicting that, in humans, PR-A mediated repression of PR-B is stronger than in mouse. We use reporter assays with human and mouse PRs to detect species differences in the repressive effects of PR-A. Results: We found that human PR-B is less sensitive to repression by human PR-A than mouse PR-B, contrary to our prediction. The difference between human and mouse PR-B sensitivity is most pronounced at PR-A/PR-B ratios typical for the preterm myometrium. Conclusions: Our results are inconsistent with the ISH. We speculate that, instead, the lower sensitivity of human PR-B to PR-A may be relevant for the maintenance of pregnancy at high progesterone levels and increasing PR-A concentrations towards term.

Disrupting the three-dimensional regulatory topology of the Pitx1 locus results in overtly normal development

ABSTRACT Developmental gene expression patterns are orchestrated by thousands of distant-acting transcriptional enhancers. However, identifying enhancers essential for the expression of their target genes has proven challenging. Maps of long-range regulatory interactions may provide the means to identify enhancers crucial for developmental gene expression. To investigate this hypothesis, we used circular chromosome conformation capture coupled with interaction maps in the mouse limb to characterize the regulatory topology of Pitx1, which is essential for hindlimb development. We identified a robust hindlimb-specific interaction between Pitx1 and a putative hindlimb-specific enhancer. To interrogate the role of this interaction in Pitx1 regulation, we used genome editing to delete this enhancer in mouse. Although deletion of the enhancer completely disrupts the interaction, Pitx1 expression in the hindlimb is only mildly affected, without any detectable compensatory interactions between the Pitx1 promoter and potentially redundant enhancers. Pitx1 enhancer null mice did not exhibit any of the characteristic morphological defects of the Pitx1−/− mutant. Our results suggest that robust, tissue-specific physical interactions at essential developmental genes have limited predictive value for identifying enhancer mutations with strong loss-of-function phenotypes. Summary: Long-range regulatory interactions may have limited predictive value for identifying enhancer mutations with strong loss-of-function phenotypes during development.