MicroRNA-196a regulates bovine newborn ovary homeobox gene (NOBOX) expression during early embryogenesis

Abstract

BackgroundOocyte-derived maternal RNAs drive early embryogenesis when the newly formed embryo is transcriptionally inactive. Recent studies in zebrafish have identified the role of microRNAs during the maternal-to-embryonic transition (MET). MicroRNAs are short RNAs that bind to the 3' UTR of target mRNAs to repress their translation and accelerate their decay. Newborn ovary homeobox gene (NOBOX) is a transcription factor that is preferentially expressed in oocytes and essential for folliculogenesis in mice. NOBOX knockout mice are infertile and lack of NOBOX disrupts expression of many germ-cell specific genes and microRNAs. We recently reported the cloning and expression of bovine NOBOX during early embryonic development and our gene knockdown studies indicate that NOBOX is a maternal effect gene essential for early embryonic development. As NOBOX is a maternal transcript critical for development and NOBOX is depleted during early embryogenesis, we hypothesized that NOBOX is targeted by microRNAs for silencing and/or degradation.ResultsUsing an algorithm "MicroInspector", a potential microRNA recognition element (MRE) for miR-196a was identified in the 3' UTR of the bovine NOBOX mRNA. Expression analysis of miR-196a in bovine oocytes and during early embryonic development indicated that it is expressed both in oocytes and embryos and tends to increase at the four-cell and eight-cell stages. Ectopic expression of NOBOX and miR-196a in HeLa cells inhibited the expression of NOBOX protein compared to the control cells without miR-196a. Similarly, the activity of a luciferase construct containing the entire 3' UTR of bovine NOBOX was suppressed, and the regulation was abolished by mutations in the miR-196a binding site indicating that the predicted MRE is critical for the direct and specific binding of miR-196a to the NOBOX mRNA. Furthermore, ectopic expression of miR-196a mimic in bovine early embryos significantly reduced the NOBOX expression at the both mRNA and protein levels.ConclusionCollectively, our results demonstrate that miR-196a is a bona fide negative regulator of NOBOX during bovine early embryogenesis.

Evaluation of MicroRNAs miR-196a, miR-30a-5P, and miR-490 as Biomarkers of Disease Activity among Patients with FSGS

“…Dysfunction and/or decrease in miRNAs of the miR-30 family could lead to podocyte apoptosis and depletion, glomerulosclerosis, and proteinuric renal disease. In other work, we found that the miR-30 family is expressed in renal tubular cells (J. Wu unpublished observations), and miR-196a was also shown to be expressed predominantly in the kidney (38). miR-196a was previously shown to correlate negatively with disease activity in SLE and has critical roles in cancer pathogenesis, viral immunity, and cell differentiation (39) as well as a role in kidney development.…”

Efectos de los RNAm maternos sobre la maduración del ovocito y el desarrollo embrionario temprano en mamíferos

“…The action of miRNAs on maternal mRNA degradation has been clear in Zebrafish embryos, which shows that the miR430 is abundant in the embryonic activation point, just at the moment the degradation of the maternal transcripts takes place (44) . Recent studies have shown that in the bovine, maternal mRNA as NOBOX and NPM2 are regulated under the action of the miR-181st and miR196a, respectively (104,105) . It is important to point out that the route of the degradation mediated by miRNAs is present only during the activation of the embryonic genome, and is absent during the oogenesis and in the initial stages of the embryo (47,52,54,106) .…”

“…La acción de miRNAs sobre la degradación de RNAm maternos ha quedado clara en embriones del Zebrafish, donde se observa que el miR430 es abundante en el punto de la activación embrionaria, justo en el momento en que sucede la degradación de los transcritos maternos (44) . Estudios recientes han demostrado que en el bovino, RNAm maternos como NOBOX y NPM2 son regulados bajo la acción de los miR-181a y miR196a, respectivamente (104,105) . Es importante señalar que la vía de degradación mediada por los of mice brg1-/-are arrested at the 2-4 cells stage (108) .…”

“…Recently it was found that the messenger of the maternal gene NOBOX has binding sites for the miR-196a in his extreme UTR 3', demonstrating that this miRNA is a negative regulator during early embryogenesis in cattle. The miR-196a is present in the oocyte and embryo, its expression is increased in the embryonic stage of 4 to 8 cells (105) . The nucleoplasmin 2 (NPM2) is a specific protein from the egg that acts on the nuclear organization in early embryonic development (80) .…”

“…Recientemente se encontró que el mensajero del gen materno NOBOX tiene sitios de unión para el miR-196a en su extremo UTR 3', demostrando que este miRNA es un regulador negativo durante la embriogénesis temprana en el bovino. El miR-196a está presente en el ovocito y en el embrión, su expresión se incrementa en el estadio embrionario de 4 y 8 células (105) . La Nucleoplasmina 2 (NPM2) es una proteína específica del ovocito que actúa en la organización nuclear en el desarrollo embrionario temprano (80) .…”