Abstract
Spatiotemporal expression of transcription factors is crucial for genomic reprogramming. Pou5f1 (Oct4) is an essential transcription factor for reprogramming. A recent study reported that OCT4A, which is crucial for establishment and maintenance of pluripotent cells, is expressed in oocytes, but maternal OCT4A is dispensable for totipotency induction. Whereas another study reported that OCT4B, which is not related to pluripotency, is predominantly expressed instead of OCT4A during early preimplantation phases in mice. To determine the expression states of OCT4 in murine preimplantation embryos, we conducted in-depth expression and functional analyses. We found that pluripotency-related OCT4 mainly localizes to the cytoplasm in early preimplantation phases, with no major nuclear localization until the 8–16-cell stage despite high expression in both oocytes and early embryos. RNA-sequencing analysis using oocytes and early preimplantation embryos could not identify the splice variants creating alternative forms of OCT4 protein. Forced expression of OCT4 in zygotes by the injection of polyadenylated mRNA clearly showed nuclear localization of OCT4 protein around 3–5-fold greater than physiological levels and impaired developmental competency in a dose-dependent manner. Embryos with modest overexpression of OCT4 could develop to the 16-cell stage; however, more than 50% of the embryos were arrested at this stage, similar to the results for OCT4 depletion. In contrast, extensive overexpression of OCT4 resulted in complete arrest at the 2-cell stage accompanied by downregulation of zygotically activated genes and repetitive elements related to the totipotent state. These results demonstrated that OCT4 protein localization was spatiotemporally altered during preimplantation development, and strict control of Oct4 protein levels was essential for proper totipotential reprogramming.
Highlights
Cellular reprogramming and differentiation are accompanied by dynamic transcriptional changes and transcription factors play central roles in both processes (Buganim et al 2013)
Oct4 expression levels differed in each stage, the coding region of the mRNA was expressed from oocyte to late preimplantation phases
At oocyte to 4-cell stages, the OCT4A staining patterns using the C10 antibody (Oct4-C10) did not show the nuclear localization typically seen in pluripotent stem cells (Fig. 1C)
Summary
Cellular reprogramming and differentiation are accompanied by dynamic transcriptional changes and transcription factors play central roles in both processes (Buganim et al 2013). Dramatic alterations of cellular fates occur during preimplantation embryo development (Burton & Torres-Padilla 2014). After fertilization, most maternal factors are degraded and zygotic gene activation (ZGA) occurs by the 2-cell stage (Hamatani et al 2004). An important transcription factor responsible for sustaining pluripotency is the POU5F1 protein ( known as OCT4) (Nichols et al 1998). Oct is crucial for driving ZGA (Lee et al 2013, Leichsenring et al 2013). Oct expression defines pluripotent stem cell fates
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