Estrogen receptor-mediated regulation of microRNA inhibits proliferation of vascular smooth muscle cells.

OBJECTIVE: Estradiol (E2) regulates gene transcription by activating estrogen receptor-α and estrogen receptor-ß. Many of the genes regulated by E2 via estrogen receptors are repressed, yet the molecular mechanisms that mediate E2-induced gene repression are currently unknown. We hypothesized that E2, acting through estrogen receptors, regulates expression of microRNAs (miRs) leading to repression of expression of specific target genes. METHODS AND RESULTS: Here, we report that E2 significantly upregulates the expression of 26 miRs and downregulates the expression of 6 miRs in mouse aorta. E2-mediated upregulation of one of these miRs, miR-203, was chosen for further study. In cultured vascular smooth muscle cells (VSMC), E2-mediated upregulation of miR-203 is mediated by estrogen receptor-α (but not estrogen receptor-ß) via transcriptional upregulation of the primary miR. We demonstrate that the transcription factors Zeb-1 and AP-1 play critical roles in mediating E2-induced upregulation of miR-203 transcription. We show further that miR-203 mediates E2-induced repression of Abl1, and p63 protein abundance in VSMC. Finally, knocking-down miR-203 abolishes E2-mediated inhibition of VSMC proliferation, and overexpression of miR-203 inhibits cultured VSMC proliferation, but not vascular endothelial cell proliferation. CONCLUSIONS: Our findings demonstrate that E2 regulates expression of miRs in the vasculature and support the estrogen receptors-dependent induction of miRs as a mechanism for E2-mediated gene repression. Furthermore, our findings demonstrate that miR-203 contributes to E2-induced inhibition of VSMC proliferation and highlight the potential of miR-203 as a therapeutic agent in the treatment of proliferative cardiovascular diseases.

Asymmetric Localization of CK2α During Xenopus Oogenesis.

The establishment of the dorso-ventral axis is a fundamental process that occurs after fertilization. Dorsal axis specification in frogs starts immediately after fertilization, and depends upon activation of Wnt/ß-catenin signaling. The protein kinase CK2α can modulate Wnt/ß-catenin signaling and is necessary for dorsal axis specification in Xenopus laevis. Our previous experiments show that CK2α transcripts and protein are animally localized in embryos, overlapping the region where Wnt/ß-catenin signaling is activated. Here we determined whether the animal localization of CK2α in the embryo is preceded by its localization in the oocyte. We found that CK2α transcripts were detected from stage I, their levels increased during oogenesis, and were animally localized as early as stage III. CK2α transcripts were translated during oogenesis and CK2α protein was localized to the animal hemisphere of stage VI oocytes. We cloned the CK2α 3'UTR and showed that the 2.8 kb CK2α transcript containing the 3'UTR was enriched during oogenesis. By injecting ectopic mRNAs, we demonstrated that both the coding and 3'UTR regions were necessary for proper CK2α transcript localization. This is the first report showing the involvement of coding and 3'UTR regions in animal transcript localization. Our findings demonstrate the pre-localization of CK2α transcript and thus, CK2α protein, in the oocyte. This may help restrict CK2α expression in preparation for dorsal axis specification.

A role for CK2alpha/beta in Xenopus early embryonic development.

CK2 is expressed widely in early embryonic development in several animal models, however its developmental role is unclear. One of the substrates of CK2 that is important in embryonic development is beta-catenin, the transcriptional co-activator of the canonical Wnt signaling pathway. This pathway has been implicated in diverse aspects of embryonic development, including one of the earliest events in embryonic development, the establishment of the dorso-ventral embryonic axis. In Xenopus laevis, dorso-ventral axis formation is dependent upon stabilization of beta-catenin in the future dorsal side of the embryo. Since CK2 phosphorylation of beta-catenin stabilizes it, we hypothesized that CK2 might be critical to upregulation of beta-catenin in Xenopus embryos and to the process of axis establishment. Our results demonstrate that CK2 is required for dorsal axis formation and is for normal upregulation of Wnt signaling genes and targets. Thus, CK2 is a regulator of endogenous axis formation in vertebrates.