Transcriptomic analysis reveals the regulation network of BmKruppel homolog 1 in the oocyte development of Bombyx mori

Kriippel homolog 1(Kr-hl),a zinc finger transcription factor,is involved in the metamorphosis and adult reproduction of insects.However,the role of Kr-hl in reproduction of holometabolic insects remains to be elucidated.The regulation network of 7-associated genes in the reproduction in Bombyx mori was investigated in this study.The higher expression level of BmKr-hl in the ovaries was detected during the late pupal stage and adults.RNA interference(RNAi)-mediated depletion of BmKr-hl in the female at day 6 of pupae resulted in abnormal oocytes at 48 h post-double-stranded RNA treatment,which showed less yolk protein deposition and partially transparent chorion.RNA-seq and subsequent differentially expressed transcripts analysis showed that knockdown of BmKr-hl caused a decrease in the expression of 2882 genes and an increase in the expression of 2565 genes in the oocytes at day 8 of pupae.Totally,27 genes coding for transcription factors were down-regulated,while six genes coding for other transcription factors were up-regulated.BmKr-hl bound to the Kr-hl binding site of the transcription factors AP-1(activating protein-1)and FOXG1 to increase their messenger RNA tran・scripts in the BmN cells,respectively.Gene Ontology enrichment and Kyoto Encyclopedia of Genes and Genomes pathway analyses of that positively co-expressed with AP-1 and FOXG1 transcripts showed mainly enrichment in the metabolic-related pathways,the nu・tri ent absorption and the yolk protein absorption processes.These data suggested that BmKr-h 1 might directly regulate the metabolic-related pathways,the nutrient absorption and the yolk protein absorption processes or probably through AP-1 and/or FOXG1 to regulate oocyte development.

Effects of MEK inhibitor U0126 on meiotic progression in mouse oocytes: microtuble organization, asymmetric division and metaphase Ⅱ arrest

In this study we used U0126, a potent and specific inhibitor of MEK, to study the roles of MEK/ERK/p90rsk signaling pathway in the meiotic cell cycle of mouse oocytes. The phosphorylation of MAP kinase and p90rsk in the oocytes treated with 1.5 μMU0126 was the same as that in oocytes cultured in drug-free medium. With 1.5 μM U0126 treatment, the spindles appeared normal as they formed in oocytes, but failed to maintain its structure.Instead, the spindle lost one pole or elongated extraordinarily. After further culture, some oocytes extruded gigantic polar bodies (＞30 μm) that later divided into two small ones. Some oocytes underwent symmetric division and produced two equal-size daughter cells in which normal spindles formed. In oocytes with different division patterns,MAP kinase was normally phosphorylated. When the concentration of U0126 was increased to 15 mM, the phosphorylation of both MAPK and p90rsk were inhibited, while symmetric division was decreased. When incubating in medium containing 15 μM U0126 for 14 h, oocytes were activated, but part of them failed to emit polar bodies. MII oocytes were also activated by 15 μM U0126, at the same time the dephosphorylation of MAP kinase and p90rsk was observed. Our results indicate that 1) MEK plays important but not indispensable roles in microtubule organization;2) MEK keeps normal meiotic spindle morphology, targets peripheral spindle positioning and regulates asymmetric division by activating some unknown substrates other than MAP kinase/p90rsk; and 3) activation of MEK/ERK/p90rsk cascade maintains MII arrest in mouse oocytes.