Temporal chromatin and transcriptome dynamics driven by JUND and progesterone receptor binding in the pregnant mouse myometrium

This study elucidates the dynamic interplay between chromatin accessibility and transcriptome remodeling in the pregnant mouse myometrium, revealing that the temporal transition from late gestation to labor is driven by stage-specific binding of JUND and progesterone receptor isoforms (PRA and PRB) that regulate gene expression and chromatin states.

The Gist

Imagine the uterus as a highly specialized factory that has one massive job: to grow a baby and then, at the perfect moment, switch gears to push that baby out, before finally cleaning up and resetting the factory for the next time.

This paper is like a detailed security camera log and blueprint analysis of that factory (specifically the muscular wall, called the myometrium) during the final days of pregnancy, the moment of birth, and the first few days after. The researchers wanted to know: How does the factory know when to stop growing, start pushing, and then clean up?

Here is the story of what they found, broken down into simple concepts:

1. The "Open Door" Policy (Chromatin Accessibility)

Think of your DNA as a massive library of instruction manuals. Usually, most of these books are locked in a vault (tightly packed chromatin) so the workers can't read them. To read a book, you have to unlock the vault door. This is called chromatin accessibility.

• The Discovery: The researchers found that just before labor starts, the factory doesn't just unlock a few doors; it blows the locks off the entire vault. Suddenly, thousands of new instruction manuals become accessible. This happens before the baby is actually born, preparing the factory for the big event.
• The Aftermath: Once the baby is born, the factory goes into "shutdown mode." It slams the vault doors shut on the "labor" instructions and starts unlocking a new set of books needed for cleaning and repair.

2. The Foremen (Transcription Factors)

The "books" (genes) don't read themselves. You need foremen (proteins called transcription factors) to open the books and tell the workers what to build. The study identified three main teams of foremen:

• Team AP-1 (The "Push" Crew): Led by a foreman named JUND.
  • Role: These guys show up right before labor. They unlock the doors to the "contraction" manuals. They work hand-in-hand with the Progesterone Receptor (PR).
  • The Twist: There are two types of PR foremen: PRB (the "Stop" sign) and PRA (the "Go" sign). During pregnancy, PRB is in charge, keeping the factory quiet. But right before labor, PRA takes over, teams up with JUND, and says, "Okay, it's time to push!"
• Team HOX (The "Cleanup" Crew):
  • Role: These foremen show up after the baby is born (postpartum). They unlock the manuals for "wound healing" and "rebuilding." They help the uterus shrink back down to its normal size.
• Team SOX and ETS:
  • Role: These are the specialized engineers who help Team AP-1 fine-tune the contractions during labor.

3. The "Silent Alarm" (Repressed Genes)

The researchers also looked at a "security system" called H3K27me3. Think of this as a heavy chain and padlock placed on specific books to make sure they never get read at the wrong time.

• The Finding: During pregnancy, the "Stop" foreman (PRB) helps put these heavy chains on the "Postpartum Cleanup" books. This ensures the uterus doesn't start shrinking and healing while the baby is still inside.
• The Release: Once the baby is born, the chains are cut (eroded), allowing the "Cleanup Crew" (HOX factors) to finally read those books and start the recovery process.

4. The "Radio Broadcast" (Enhancer RNAs)

The researchers also listened for "radio broadcasts" coming from the unlocked doors. These are short signals called eRNAs.

• They found that right before and during labor, the factory is buzzing with these radio signals, confirming that the "Push" crew is actively reading the manuals and getting ready to work.

The Big Picture Analogy

Imagine a theater production:

1. Late Pregnancy (D17): The stage is set, but the actors are resting. The "Stop" sign (PRB) is up.
2. Pre-Labor (PL): The "Go" sign (PRA) teams up with the Director (JUND). They unlock the script (chromatin opens up). The actors (genes) start rehearsing.
3. Labor (LAB): The show is on! The "Push" crew is in full swing. The "Stop" sign is gone.
4. Postpartum (D4PP): The show is over. The stagehands (HOX factors) come in, unlock the "Cleanup" scripts, and start dismantling the set to reset the theater for the next show.

Why Does This Matter?

Understanding exactly who unlocks the doors and when helps scientists figure out what goes wrong in difficult pregnancies.

• If the "Push" crew doesn't show up, you might get preterm labor (the show starts too early).
• If the "Stop" sign never comes down, you might get postpartum hemorrhage (the factory can't shut down and clean up properly).

This study gives us a new, detailed map of the molecular switches that control birth, offering potential new ways to fix these switches if they get stuck.

Technical Summary

1. Problem Statement

The myometrium (uterine muscle) undergoes profound phenotypic transitions from a quiescent state during pregnancy to a highly contractile state during labor, followed by involution (regression) postpartum. While transcriptomic changes associated with these stages are well-documented, the underlying epigenetic mechanisms driving these transitions remain poorly characterized.

• The Gap: Previous studies indicated that active histone marks (H3K27ac, H3K4me3) do not significantly change between pregnancy and labor, suggesting that other regulatory layers control the transition.
• The Question: How do chromatin accessibility dynamics, transcription factor (TF) binding, and repressive histone modifications coordinate to regulate the myometrial shift from quiescence to contraction and back to a non-pregnant state?

2. Methodology

The study utilized a multi-omics approach on mouse myometrium tissues collected at specific gestational time points:

• Time Points: Late gestation (Day 15, Day 17), Prelabor (Day 19, non-laboring), Term Labor (Day 19.5), and Postpartum (Day 1 and Day 4).
• Techniques:
  • ATAC-seq: To map genome-wide chromatin accessibility and identify transcription factor binding footprints.
  • RNA-seq (Total): To analyze both exon and intron reads, capturing primary transcript levels and nascent transcription.
  • ChIP-seq: Performed for:
    • JUND: An AP-1 family transcription factor.
    • Progesterone Receptor (PR): Both Total PR (tPR, recognizing PRA and PRB) and the specific PRB isoform.
    • H3K27me3: A repressive histone mark.
  • Bioinformatics: Differential accessibility/expression analysis (DiffBind, DESeq2), motif enrichment (TOBIAS), and integrative analysis of enhancer RNAs (eRNAs).

3. Key Contributions

This study provides the first comprehensive, genome-wide temporal map of chromatin accessibility and TF binding dynamics in the mouse myometrium across the entire pregnancy-labor-postpartum continuum. Key contributions include:

1. Discovery of Pre-labor Chromatin Opening: Identification of a massive global increase in chromatin accessibility before the onset of active labor contractions.
2. Isoform-Specific PR Dynamics: Differentiation between the roles of PR isoforms (PRA vs. PRB) in regulating labor genes versus repressing postpartum genes.
3. Enhancer Identification: Discovery of labor-specific enhancers producing eRNAs, driven by a cooperative network of AP-1, SOX, and ETS transcription factors.
4. Postpartum Involution Mechanism: Elucidation of how HOX transcription factors and H3K27me3 erosion drive the reprogramming of the myometrium back to a non-pregnant state.

4. Detailed Results

A. Chromatin Accessibility (ATAC-seq)

• Global Opening: There is a dramatic increase in chromatin accessibility (~19,000 regions) between Day 17 and the prelabor stage (Day 19).
• Labor vs. Postpartum: A massive "closing" of chromatin occurs immediately after labor (Day 1 postpartum), followed by the opening of new regions (~36,000) by Day 4 postpartum.
• Motif Footprinting:
  • Prelabor/Labor: Enriched for AP-1 (specifically JUND), SOX, and ETS family motifs.
  • Pregnancy (D17): Enriched for Progesterone Receptor (PR) and CTCF motifs.
  • Postpartum (D4PP): Enriched for HOX family motifs.

B. Transcriptomics (RNA-seq)

• Pre-labor Surge: The most significant transcriptional upregulation occurs at the prelabor stage (D19), not during active labor. This includes contraction-associated genes (Oxtr, Gja1, Mmp8/11/28) and inflammatory chemokines.
• Labor Transition: The transition from prelabor to active labor is characterized primarily by a downregulation of many genes, suggesting the machinery is already primed.
• Postpartum: Distinct upregulation of ECM remodeling genes and HOX family genes (Hoxa9-11, Hoxd9-11) involved in tissue reorganization.

C. Transcription Factor Binding (ChIP-seq)

• JUND (AP-1):
  • Binds to accessible regions as early as Day 17 ("pre-marking").
  • Binding increases significantly at prelabor and labor.
  • Binding is lost by Day 4 postpartum.
• Progesterone Receptor (PR) Isoforms:
  • PRA (Total PR without PRB): Co-binds with JUND at prelabor/labor accessible regions (e.g., Gja1 promoter), acting as an activator.
  • PRB: Predominantly binds during pregnancy (D17) and postpartum (D4PP).
    • Repressive Role: PRB is found at H3K27me3 repressed regions.
    • Pre-marking: PRB binds to regions that are later repressed by H3K27me3 at the prelabor stage, suggesting a mechanism to silence postpartum genes during pregnancy.
    • Absence in Labor: PRB is strikingly absent from labor-associated accessible chromatin.

D. Enhancer Activity (eRNAs)

• Analysis of intergenic accessible regions revealed 100 candidate labor enhancers producing elevated eRNAs at prelabor/labor.
• These enhancers are enriched for AP-1, SOX, and ETS motifs.
• A specific enhancer downstream of the Fos gene was identified as conserved across mouse, rat, and human, bound by JUND and PR, and active during labor.

E. Repressive Mechanisms (H3K27me3)

• H3K27me3 dynamics did not correlate with chromatin accessibility changes (no "poised" enhancers found).
• Instead, H3K27me3 is used to repress postpartum-specific genes (like Hoxd cluster) during pregnancy.
• Erosion: At Day 4 postpartum, H3K27me3 erodes over the Hoxd locus, allowing the expression of Hoxd genes required for involution.

5. Significance and Conclusion

This study redefines the molecular timeline of labor initiation:

1. Pre-labor Priming: The myometrium undergoes a global chromatin opening and transcriptional surge before labor begins, driven by JUND and PRA (not PRB).
2. Isoform Switching: The transition to labor involves a switch from PRB-mediated repression (during pregnancy) to PRA/JUND-mediated activation.
3. New Regulatory Networks: It identifies SOX and ETS factors as critical co-regulators with AP-1 in the labor network.
4. Postpartum Recovery: It reveals a distinct mechanism for uterine involution involving HOX factors and the removal of repressive H3K27me3 marks.

Clinical Implication: These findings provide new therapeutic targets for reproductive disorders. Understanding the specific roles of PR isoforms and AP-1 complexes could lead to interventions for preterm labor (by modulating the premature opening of chromatin) or postpartum hemorrhage (by understanding the failure of involution mechanisms).