cGMP-dependent ADP depolymerization of actin mediates estrogen increase in cervical epithelial permeability

Abstract

Estrogen increases secretion of cervical mucus in women, and the effect depends on fragmentation of the cytoskeleton. The objective of the present study was to understand the molecular mechanism of estrogen action. Treatment of human cervical epithelial cells with 17beta-estradiol, sodium nitroprusside (SNP), or 8-bromoguanosine 3', 5'-cyclic monophosphate (8-Br-cGMP) increased cellular monomeric G-actin and decreased polymerized F-actin. The effects of estradiol were blocked by tamoxifen, by the guanylate cyclase inhibitor LY-83583, and by the cGMP-dependent protein kinase inhibitor KT-5823. The effects of SNP were blocked by LY-83583 and KT-5823, while the effects of 8-Br-cGMP were blocked only by KT-5823. Treatment with phalloidin decreased paracellular permeability and G-actin. Treatment with 17beta-estradiol, SNP, or 8-Br-cGMP attenuated SNP-induced phosphorylation of [(32)P]adenylate NAD in vitro: tamoxifen blocked the effect of estrogen; LY-83583 blocked the effect of SNP but not that of 8-Br-cGMP, while KT-5823 blocked effects of both SNP and 8-Br-cGMP. These results indicate that estrogen, nitric oxide (NO), and cGMP stimulate actin depolymerization. A possible mechanism is NO-induced, cGMP-dependent protein kinase augmentation of ADP-ribosylation of monomeric actin.

Non-Muscle Myosin-II-B Filament Regulation of Paracellular Resistance in Cervical Epithelial Cells Is Associated With Modulation of the Cortical Acto-Myosin

“…1, Table 1). These results confirm previous studies [4,5] that modulation of actin polymerization affects the paracellular permeability.…”

“…In those experiments purified NMM-II-B filaments were exposed to 0.2 μg per 100 μl of purified CK2 from rat liver (Sigma, dissolved in PBS), or to heat-inactivated (65°C) CK2. Cellular G-actin content was determined by the DNase-I Inhibition Assay as previously described [4,5]. Briefly, cells on filters were lysed in situ and DNase-I activity in the lysate was assayed by measuring DNase-I-dependent degradation of DNA.…”

“…F-actin) [30]. To test whether in the epithelial CaSki cells actin polymerization status affects NMM-II-B filamentation, the above experiments were repeated using preparations obtained from cells treated with phalloidin, to stimulate actin polymerization [5], or with sodium nitroprusside (SNP), to stimulate actin depolymerization [4]. The results in Tables 2 and 3 show that the assembly and disassembly rates of NMM-II-B filaments were not significantly affected by prior treatments with phalloidin or SNP.…”

“…The actin cytoskeleton can be modulated by regulation of actin polymerization. Formation of G-actindominated cytoskeleton decreases the R LIS , while actin polymerization (F-actin) produces rigid cytoskeleton and increases R LIS [4,5].…”

Luminal flow induces eNOS activation and translocation in the rat thick ascending limb. II. Role of PI3-kinase and Hsp90

“…In the brain, eNOS has been localized to the ciliated epithelium of the ependymal cells lining the ventricles (58). In the testis, eNOS is localized to the seminiferous tubules and Leydig cells (58,60), where its function is still unclear, whereas eNOS expressed in cervical epithelial cells is known to increase paracellular permeability and mucus secretion (21)(22)(23). To our knowledge, there are no data regarding regulation of eNOS in any of these cells; nevertheless, given that they are subjected to changes in luminal flow or mechanical stimulation, it is likely that PI3-kinase-dependent eNOS translocation and activation play a role in the physiology of these cells.…”