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Oct4 gene key to human embryo development
For the first time, researchers have used genome editing tech to reveal the role of a key gene in a human embryo. Their hope is that by identifying specific gene functions, they will better understand the biology of early development and thus affect better IVF treatment. The work was published in Nature.
The team was led by scientists at the Francis Crick Institute collaborating with colleagues at Cambridge University, Oxford University, the Wellcome Trust Sanger Institute, Seoul National University and Bourn Hall Fertility Clinic in Cambridge, United Kingdom (UK). The work was chiefly funded by the UK Medical Research Council.
Wellcome and Cancer Research UK used genome editing techniques, known as CRISPR-Cas9 to stop a key gene from producing a protein called OCT4, which normally becomes active in the first few days of human embryo development. After the egg is fertilized, it continually divides until it forms a ball of around 200 cells called a blastocyst. Researchers were able to see that the human embryo must have OCT4 in order to correctly form into a blastocyst!
"We were surprised to see just how crucial this gene is for human embryo development, but we need to continue our work to confirm its role. Other research methods, including studies in mice, suggest a later and more focused role for OCT4, so our results highlight the need for more human embryo research."
"One way to find out what a gene does in the developing embryo is to see what happens when it isn't working," explains Kathy K. Niakan PhD, of the Francis Crick Institute, who led the research. "Now that we have an efficient way of doing this, we hope other scientists will use it to find out the roles of other genes. If we know the key genes that embryos need to develop successfully, we could improve IVF treatments and understand some causes of pregnancy failure. It may take many years to achieve such an understanding — our study is just the first step."
The team spent over a year optimising their techniques using mouse embryos and human embryonic stem cells before starting work on human embryos. To inactivate OCT4, they used CRISPR/Cas9 to change the DNA of 41 human embryos. After seven days, embryo development was stopped and the embryos were analyzed. Embryos used in the study were donated by couples who had undergone IVF treatment and had frozen embryos remaining in storage.
The majority of embryos used were donated by couples who had completed their family, and wanted their surplus embryos used for research. The study was done under strict regulatory oversight by the Human Fertilization and Embryology Authority (HFEA), the UK Government's independent regulator overseeing infertility treatment and research.
As well as in human embryo development, OCT4 is thought to be important in stem cell biology. 'Pluripotent' stem cells can become any type of body cell — and they can be derived from embryos or created from adult cells such as skin cells. Human embryonic stem cells are taken from the developing embryo with high levels of OCT4.
"We have the technology to create and use pluripotent stem cells, which is undoubtedly a fantastic achievement, but we still don't understand exactly how these cells work," explains Dr James Turner, co-author of the study from the Francis Crick Institute. "Learning more about how different genes cause cells to become and remain pluripotent will help us to produce and use stem cells more reliably."
According to Kay Elder PhD, study co-author, the Bourn Hall Clinic: "Successful IVF treatment is crucially dependent on culture systems that provide an optimal environment for healthy embryo development. Many embryos arrest in culture, or fail to continue developing after implantation; this research will significantly help treatment for infertile couples, by helping us to identify the factors that are essential for ensuring that human embryos can develop into healthy babies."
Ludovic Vallier PhD, study co-author from the Wellcome Trust Sanger Institute and the Wellcome - MRC Cambridge Stem Cell Institute, adds: "The acquisition of this knowledge will be essential to develop new treatments against developmental disorders and could also help us understand adult diseases such as diabetes that may originate during the early stage of life. Thus, this research will open new fields of opportunity for basic and translational medicine applications."
Despite their fundamental biological and clinical importance, the molecular mechanisms that regulate the first cell fate decisions in the human embryo are not well understood. Here we use CRISPR–Cas9-mediated genome editing to investigate the function of the pluripotency transcription factor OCT4 during human embryogenesis. We identified an efficient OCT4-targeting guide RNA using an inducible human embryonic stem cell-based system and microinjection of mouse zygotes. Using these refined methods, we efficiently and specifically targeted the gene encoding OCT4 (POU5F1) in diploid human zygotes and found that blastocyst development was compromised. Transcriptomics analysis revealed that, in POU5F1-null cells, gene expression was downregulated not only for extra-embryonic trophectoderm genes, such as CDX2, but also for regulators of the pluripotent epiblast, including NANOG. By contrast, Pou5f1-null mouse embryos maintained the expression of orthologous genes, and blastocyst development was established, but maintenance was compromised. We conclude that CRISPR–Cas9-mediated genome editing is a powerful method for investigating gene function in the context of human development.
All authors: Norah M. E. Fogarty, Afshan McCarthy, Kirsten E. Snijders, Benjamin E. Powell, Nada Kubikova, Paul Blakeley, Rebecca Lea, Kay Elder, Sissy E. Wamaitha, Daesik Kim, Valdone Maciulyte, Jens Kleinjung, Jin-Soo Kim, Dagan Wells, Ludovic Vallier, Alessandro Bertero, James M. A. Turner & Kathy K. Niakan
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CRISPR-Cas9 used to cut out the protein Oct4 in (LEFT) normally dividing human embryo — during the first 200 cell divisions — reveals how the orderly cells become disorganized in the (RIGHT) resulting embryo. Image Credit: Francis Crick Institute, London, UK