The Role of Oct4 in Stem Cells

The Role of Oct4 in Stem Cells

Instructor:

Focus:

I selected a group of papers that all relate to the study of transcriptional regulation of embryonic stem cells. The connection among the articles is not focused on an individual’s research program; instead I chose to focus the module on understanding the function of a single transcription factor (Oct4) and its role as a regulator of stem cell populations. The progression of module moves from understanding Oct4 function in the embryo to how it regulates pluripotency and affects downstream targets for maintaining a stem cell state.

Overview:

There are three papers that are part of this article. The articles range in publication date from 1998 through 2006. I chose the first paper as the starting point of the module because it details key experiments that demonstrate the requirement for Oct4 function in maintaining embryonic cells in a stem cell state. I chose second paper for the module because it shows progression towards understanding how Oct4 acts as a transcription factor and describes experiments based on quantitative techniques for measuring gene/protein activity. The final paper in the module was chosen because it represents how advanced methods in molecular biology are being employed to study gene regulation in a global manner.

Applicable for Courses:

Upper level seminar, developmental biology, cell biology

Educational Level:

Introductory

Roadmap Objectives:

    • Article: Formation of Pluripotent Stem Cells in the Mammalian Embryo Depends on the POU Transcription Factor Oct4. (1998) Nichols et al. Cell 95, 379 – 391
    • Content area/major concepts: general mammalian development, cell biology (cell- signaling, transcriptional regulator), cell cycle, homologous recombination, necessity versus sufficiency of gene function
    • Methods or technology used to obtain data: homologous recombination, targeted knock out technology, Mendelian genetics, PCR, genotyping
    • How the CREATE strategy was used: all tools were used and we discussed the article fully, including a grant panel at the end.
    • Biggest teaching challenge: complicated experimental paradigm, jargon of genetics and molecular biology, substantial number of experiments
    • Article: Quantitative expression of Oct-3/4 defines differentiation, dedifferentiation or self-renewal of ES cells. (2000) Hitoshi Niwa, Jun-ichi Miyazaki, & Austin G. Smith, Nature Genetics, 24, 372-376
    • Content area/major concepts: recombinant DNA technology, quantitative versus qualitative experiments, transcriptional regulation, RNA, DNA, antibodies, hybridization, hydrogen bonding, enzyme kinetics
    • Methods or technology used to obtain data: cell culture assay, molecular biology, conditional gene expression, Northern blot analysis, in situ hybridization, RT-PCR
    • How the CREATE strategy was used: all tools were used and we discussed the article fully, including a grant panel at the end.
    • Biggest teaching challenge: Complicated techniques, quantitative data sets
    • Article: Article 3
    • Content area/major concepts: genomics, antisense RNA, DNA binding sites, transcriptional repression/activation, role of other proteins in regulating pluripotency (Nanog)
    • Methods or technology used to obtain data: microarray technology, RNAi, chromatin immunoprecipitation, quantitative DNA analysis…and other related methods covered in previous papers
    • How the CREATE strategy was used: all tools were used and we discussed the article fully, including a grant panel at the end.
    • Biggest teaching challenge: difficulty with genomics, complicated data sets, accessibility to scientists involved in project

Advice for Using Module/Activity:

Use supplemental data as a way to assess understanding of discussions.
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