Transposon-mediated rewiring of gene regulatory networks contributed to the evolution of pregnancy in mammals

Journal name:
Nature Genetics
Volume:
43,
Pages:
1154–1159
Year published:
DOI:
doi:10.1038/ng.917
Received
Accepted
Published online

A fundamental challenge in biology is explaining the origin of novel phenotypic characters such as new cell types1, 2, 3, 4; the molecular mechanisms that give rise to novelties are unclear5, 6, 7. We explored the gene regulatory landscape of mammalian endometrial cells using comparative RNA-Seq and found that 1,532 genes were recruited into endometrial expression in placental mammals, indicating that the evolution of pregnancy was associated with a large-scale rewiring of the gene regulatory network. About 13% of recruited genes are within 200 kb of a Eutherian-specific transposable element (MER20). These transposons have the epigenetic signatures of enhancers, insulators and repressors, directly bind transcription factors essential for pregnancy and coordinately regulate gene expression in response to progesterone and cAMP. We conclude that the transposable element, MER20, contributed to the origin of a novel gene regulatory network dedicated to pregnancy in placental mammals, particularly by recruiting the cAMP signaling pathway into endometrial stromal cells.

At a glance

Figures

  1. Evolution of the endometrial stromal cell transcriptome in Therian mammals.
    Figure 1: Evolution of the endometrial stromal cell transcriptome in Therian mammals.

    (a) Amniote phylogeny showing approximate divergence dates between major lineages; opossum, armadillo and human samples were included in this study. Placental mammals are indicated in red. (b) Venn diagram showing the intersection of 1:1:1 homologous genes expressed in endometrial cells of human, armadillo and opossum inferred from RNA-Seq. In total, 1,532 genes were scored as expressed in both human and armadillo but not opossum.

  2. MER20s are over-represented near progesterone/cAMP-responsive endometrial genes and have genomic and epigenetic signatures of regulatory elements.
    Figure 2: MER20s are over-represented near progesterone/cAMP-responsive endometrial genes and have genomic and epigenetic signatures of regulatory elements.

    (a) Distribution of distances from differentially regulated stromal genes (N = 6,504) to MER20s in 5-kb bins. Gray bars indicate the total number of MER20s in each bin, and brown bars indicate the distance of the closest MER20 to the gene. The number of genes with MER20s located between transcriptional start and end sites is indicated by 0. The expected number of MER20-associated genes per bin given random positions in the human genome (black line) and compared to genes that were not differentially regulated upon decidualization (blue line) are shown for the location of the closest MER20 to stromally regulated genes (mean ± s.d.). (b) MER20s are located in regions of the genome with high CpG island density, PhastCons scores and 7× regulatory potential (RP). The profile of histone modifications around MER20s located within 200 kb of genes either up- or downregulated upon differentiation of human ESCs is shown for several methylation and acetylation events and for the vertebrate insulator protein CTCF. Panel names are colored with respect to the profile shown below. MER20s are centered at position 0 (red box), with normalized ChIP-Seq tag density in 5 bp windows upstream and downstream of the MER20 shown as lines. (c) Venn diagram showing intersections among MER20s classified by histone modifications as repressors, insulators or enhancers.

  3. MER20s have binding sites for numerous transcription factors, cofactors and insulator proteins and evolve under functional constraints.
    Figure 3: MER20s have binding sites for numerous transcription factors, cofactors and insulator proteins and evolve under functional constraints.

    (a) The consensus MER20 contains putative binding sites for numerous transcription factors; only sites with a core match of greater than 0.88 are shown. Overlaid plot shows the 3-bp moving average of the per nucleotide substitution rate from a random sample of 500 MER20s. (b) Nucleotide substitution rates (per 109 years) for various classes of sequence are shown with increasing functional constraint from top to bottom (log scale). Nucleotide substitution rates of putative transcription factor binding sites (pTFBS) and non-binding sites (nonTFBS) from a are shown in red. Substitution rates for non-MER20 sequences are shown36.

  4. MER20s are bound by transcription factors and cofactors important for decidualization and pregnancy.
    Figure 4: MER20s are bound by transcription factors and cofactors important for decidualization and pregnancy.

    (a) Heat map of ChIP-qPCR data showing fold enrichment of target over normal IgG controls after normalization to input DNA (Enrich.). MER20s are named by their nearest gene. Five MER20s were enriched (>2-fold over background) for FOXO1A, PGR and C/EBPβ, 7 for HoxA-11, 8 for PRMT1/4, 9 for USF1, 10 for p300 and 15 for YY1 and CTCF. (b) Pairwise Pearson's correlation coefficients (PCCs) calculated for transcription factor binding to MER20s indicates that transcription factors with insulator functions (blue branches) coordinately bind MER20s to the exclusion of transcription factors with enhancer and/or repressor functions (yellow branches) and vice versa. (c) PCCs indicate that MER20s fall into two distinct groups based on the combination of transcription factors they bind: 'insulator-type' MER20s shown with blue branches and 'enhancer/repressor-type' with yellow branches.

  5. MER20 reporter constructs regulate luciferase expression.
    Figure 5: MER20 reporter constructs regulate luciferase expression.

    (a) Heat map shows fold changes in luciferase expression between progesterone/cAMP-treated cells and untreated cells transiently transfected with MER20 reporter constructs. Cell types are derived from mammalian cervix (HeLa), lung (A549), kidney (COS-1), muscle (MyoM), keratinocytes (PAM212), chondrocytes (CHON) and endometrial stromal cells (ESC) and chicken fibroblasts (GgaF). (b) Regulatory strength of MER20s across cell types. Values show the sum of fold changes in luciferase expression upon progesterone/cAMP treatment from Figure 4a. The greatest regulatory strength was observed for ESC, whereas MER20s had only weak regulatory ability in other cell types. (c) Expression of transcription factors shown to bind MER20s by ChIP across human tissues. The only tissue that coexpresses all transcription factors and cofactors shown to bind MER20s is the uterus.

  6. MER20s are candidate insulator elements.
    Figure 6: MER20s are candidate insulator elements.

    (a) Insulator-type MER20s are located between differentially expressed genes in human ESC. Cartoon shows the relative locations of genes (named rectangles) and MER20s (small blue or yellow rectangles). The color of each rectangle shows the fold change in expression of that gene upon progesterone/cAMP stimulation in human ESCs (green, downregulation; red, upregulation). White boxes indicate genes not expressed in human ESC. Blue and yellow boxes between genes indicate insulator-type and cis-regulatory–type MER20s, respectively. Black boxes are MER20s that were not characterized in this study. Insulator-type MER20s are significantly more common between differentially expressed genes than expected by chance (P = 0.001, binomial test). Asterisks (*) indicate MER20s that have been previously identified as regulatory elements. (b) Model of gene regulatory rewiring by MER20s. Ancestrally, numerous genes (black arrows) were not expressed in ESCs because they were repressed by epigenetic modifications of chromatin and direct silencing by transcriptional repressors. MER20s inserted into the genome in the placental mammal lineage (blue/yellow box on phylogeny), which prevented the spread of silent chromatin, establishing new borders between transcriptionally silent (green) and active (red) chromatin.

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Author information

Affiliations

  1. Department of Ecology and Evolutionary Biology & Yale Systems Biology Institute, Yale University, New Haven, Connecticut, USA.

    • Vincent J Lynch,
    • Robert D Leclerc,
    • Gemma May &
    • Günter P Wagner

Contributions

V.J.L. and G.P.W. designed experiments and wrote the manuscript. V.J.L. and G.M. performed experiments and analyzed data, and R.D.L. designed and performed bioinformatics analyses.

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The authors declare no competing financial interests.

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