Localization of regulatory protein binding sites in the proximal region of human myometrial connexin 43 gene

Abstract

Parturition is preceded by a large increase in gap junctions between myometrial smooth muscle cells. Connexin 43 is the major structural protein of myometrial gap junctions. To explore transcriptional regulation of the myometrial Cx43 gene, we used DNase I footprinting, electrophoretic mobility shift and transient transfection assays to examine a 312 bp promoter region (-164 to +148) of the gene, utilizing human myometrial cell cultures and nuclear extracts. The DNase I studies showed four regions of nucleoprotein interactions. Protection of region 1 (-80 to -31) encompassed an Activator Protein 1 (AP1) (-44 to -36) and two Specificity Protein 1 (Sp1) (-77 to -69 and -59 to -48) consensus sequences. Regions 2 to 4 included the transcription initiation site (-10 to +25), an Ets/NF-kB consensus sequence (+47 to +74) and a TA-rich region (+81 to +101) respectively. Gel mobility shift and supershift assays demonstrated c-Jun and Sp1 binding at the AP1 and Sp1 sites respectively. Promoter mutagenesis and transient transfection analyses combined with Sp1 and c-Jun/c-Fos over-expression studies indicate that both Sp1 and c-Jun are required for maximal promoter activity and, therefore, may positively regulate transcription of myometrial Cx43 during the initiation of labour.

Prostaglandin F2α regulates the expression of uterine activation proteins via multiple signalling pathways

“…The promoter region of CX43 contains NFkB binding sites, suggesting that NFkB activation can stimulate CX43 expression (Echetebu et al 1999). In this study, we found that NFkB signalling was involved in CX43 expression.…”

c-Jun N-Terminal Kinase Activation Mediates Downregulation of Connexin43 in Cardiomyocytes

“…Interestingly, activation of activator protein-1, a well characterized downstream target of JNK, has been shown to be necessary for maximal Cx43 promoter activity in human myometrial smooth muscle. 39 In addition, aortic smooth muscle cells have been shown to significantly increase steady-state levels of Cx43 mRNA in response to hypertension, 40 a known activator of JNK. 41 Together, these experiments suggest that the Cx43 promoter is subject to JNK-mediated regulation in a cell-type-specific manner.…”

Cyclic Mechanical Stretch Induces Cardiomyocyte Orientation and Polarization of the Gap Junction Protein Connexin43

“…22,23 Therefore, we wanted to examine if stretch induces nuclear accumulation of these transcription factors. In our EMSA experiments enhanced localization of both factors in the nuclei of stretched cells could clearly be demonstrated and, because previous studies of our working group indicated a dependence of Cx43 expression on nuclear presence of AP1 and CREB after ERK1/2 activation by isoprenaline, 33 it might be possible that these transcription factors are upregulated following ERK1/2 activation by stretch.…”