Abnormal uterine activity in pregnancy causes a range of important clinical disorders, including preterm birth, dysfunctional labour and post-partum haemorrhage. Uterine contractile patterns are controlled by the generation of complex electrical signals at the myometrial smooth muscle plasma membrane. To identify novel targets to treat conditions associated with uterine dysfunction, we undertook a genome-wide screen of potassium channels that are enriched in myometrial smooth muscle. Computational modelling identified Kir7.1 as potentially important in regulating uterine excitability during pregnancy. We demonstrate Kir7.1 current hyper-polarizes uterine myocytes and promotes quiescence during gestation. Labour is associated with a decline, but not loss, of Kir7.1 expression. Knockdown of Kir7.1 by lentiviral expression of miRNA was sufficient to increase uterine contractile force and duration significantly. Conversely, overexpression of Kir7.1 inhibited uterine contractility. Finally, we demonstrate that the Kir7.1 inhibitor VU590 as well as novel derivative compounds induces profound, long-lasting contractions in mouse and human myometrium; the activity of these inhibitors exceeds that of other uterotonic drugs. We conclude Kir7.1 regulates the transition from quiescence to contractions in the pregnant uterus and may be a target for therapies to control uterine contractility.

SUMMARYUbiquitination is essential for protein degradation and signaling and pivotal to many physiological processes. Ubiquitination of a subset of G-protein-coupled receptors (GPCRs) by the E3 ligase NEDD4–2 is required for p38 activation, but how GPCRs activate NEDD4–2 to promote ubiquitinmediated signaling is not known. Here, we report that the GPCR protease-activated receptor-1 (PAR1) stimulates c-Src-mediated tyrosine phosphorylation and activation of NEDD4–2 to promote p38 signaling and endothelial barrier disruption. Using mass spectrometry, we identified a unique phosphorylated tyrosine (Y)-485 within the 2,3-linker peptide between WW domain 2 and 3 of NEDD4–2 in agonist-stimulated cells. Mutation of NEDD4–2 Y485 impaired E3 ligase activity and failed to rescue PAR1-stimulated p38 activation and endothelial barrier permeability. The purinergic P2Y1 receptor also required c-Src and NEDD4–2 tyrosine phosphorylation for p38 activation. These studies reveal a novel role for c-Src in GPCR-induced NEDD4–2 activation, which is critical for driving ubiquitin-mediated p38 inflammatory signaling.

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Phosphorylation at distinct sites determines the effect of the chaperone protein HSP27 on endothelial barrier permeability.

Despite some significant differences in the first-trimester serum levels of angiogenic factors, our models suggest that these factors are not clinically useful for prediction in women who later developed PE.