MYB and ELF3 differentially modulate labor-inducing gene expression in myometrial cells

Abstract

Spontaneous uterine contractions are initiated when smooth muscle cells (SMCs) within the uterine muscle, or myometrium, transition from a functionally dormant to an actively contractile phenotype at the end of the pregnancy period. We know that this process is accompanied by gestational time point-specific differences in the SMC transcriptome, which can be modulated by the activator protein 1 (AP-1), nuclear factor kappa beta (NF-κβ), estrogen receptor (ER), and progesterone receptor (PR) transcription factors. Less is known, however, about the additional proteins that might assist these factors in conferring the transcriptional changes observed at labor onset. Here, we present functional evidence for the roles of two proteins previously understudied in the SMC context—MYB and ELF3—which can contribute to the regulation of labor-driving gene transcription. We show that the MYB and ELF3 genes exhibit elevated transcript expression levels in mouse and human myometrial tissues during spontaneous term labor. The expression of both genes was also significantly increased in mouse myometrium during preterm labor induced by the progesterone antagonist mifepristone (RU486), but not during infection-simulating preterm labor induced by intrauterine infusion of lipopolysaccharide (LPS). Furthermore, both MYB and ELF3 proteins affect labor-driving gene promoter activity, although in surprisingly opposing ways: Gja1 and Fos promoter activation increases in the presence of MYB and decreases in the presence of ELF3. Collectively, our study adds to the current understanding of the transcription factor network that defines the transcriptomes of SMCs during late gestation and implicates two new players in the control of labor timing.

SOX4 exerts contrasting regulatory effects on labor-associated gene promoters in myometrial cells

“…We assessed whether the transactivation potential of the AP-1 dimer combinations on the Gja1 and Fos gene promoters is altered in the presence of SOX4 (Fig 4A/B). Note that the two controls ( Gja1 and Fos promoter activities indicated by asterisks in Fig 4A/B) increased with the addition of JUND, and were significantly elevated in the presence of JUND and FOSL2 proteins, as previously reported (8). Here we show that the addition of SOX4 does not significantly activate the Gja1 promoter, either on its own or in association with the AP-1 proteins, despite the presence of a strong SOX4 motif in the promoter sequence (Fig 4A).…”

“…Significantly different results (p<0.05) are indicated by groups labeled with different letters. Asterisk (*) in (A) and (B) indicates control groups as previously reported in (8).…”

“…The SOX4-FLAG construct was obtained from Addgene (catalog #110360). Expression of these constructs was confirmed by western blot (see SFig2 in (8)].…”

SOX4 exerts contrasting regulatory effects on labor-associated gene promoters in myometrial cells

“…Having established that murine myometrium display increased levels of SOX4 protein at the onset of both term and preterm labor, we next sought to ascertain whether SOX4 exerts a functional role in the transcriptional control of labor-associated genes which contain strong binding motifs for both AP-1 and SOX4. We used a luciferase reporter assay as a means of measuring promoter regulation in transfected Syrian hamster myometrium (SHM) cells [ 8 , 13 , 17 ]. We used previously generated luciferase constructs with the reporter gene under the control of the well-studied labor-associated promoters from the Gja1 and Fos genes [ 8 ] to assess the regulatory capacity of SOX4, either on its own or alongside AP-1 factors.…”

“…Recently, we found that two additional factors, MYB and ELF3, exhibit differential expression in pre-laboring and laboring human and murine myometrium as SMCs undergo the quiescent-to-contractile transition. We found that despite being upregulated at labor onset, these factors play opposing roles in stimulating expression from labor-associated gene promoters, with MYB having an activating and ELF3 a repressive role [ 8 ].…”