Polo Lab: Epigenetic Processes in Pluripotency and Reprogramming

Polo Lab

Mouse embryonic stem cells stained for OCT (yellow), NANOG (magenta) and DNS (cyan)

Our research

Epigenetics in development

All cells come from a common pluripotent precursor cell and become specialised into mature cell types by tightly regulating which genes are turned on and off during development. Turning on the correct genes during the correct time of development is fundamental for proper cell maturation. More importantly, genes that are responsible for telling cells to grow and proliferate are functionally silenced once the cell matures.

In 2007 Shinya Yamanaka and colleagues showed for the first time that this program of maturing a cell can be reversed by re-expressing genes important during the pluripotent stage, Oct4, Sox2, c-Myc and Klf4. In a process termed “reprogramming” a fully mature cell was reverted to a precursor cell, an induced pluripotent stem cell (iPSC). This iPSC was able to once again divide and multiply and regained its potential to become any cell in the body.

This revolutionised the way that we think about development and diseases. With this technology we can now generate pluripotent cells from cells that are easily obtained from patients such as from skin or blood. We can then use these cells to test potential treatments or one day in the future generate cell therapies.

Although this technology has immense potential, the molecular mechanisms that govern this process are still mostly unknown. Developing a comprehensive understanding of these processes is essential in order to both understand the critical roles epigenetics plays in cell differentiation and development, and to successfully remedy its disruption in disease.

Epigenetics in cancer

It has long been known that epigenetics plays a major role in the development and progression of many cancers.  Epigenetic modifications on oncogenes and tumour suppressor genes can result in uncontrolled growth and division of cells, and ultimately cancer progression. Several cancers have been shown to reactivate the pluripotency genes OCT4, SOX2 and/or NANOG, with high expression levels positively correlating with cancer progression and severity.

Highlighted publications

  • Xiaodong Liu, Jia Ping Tan, Jan Schröder, Asma Aberkane & John F. Ouyang et al. 2021, ‘Modelling human blastocysts by re-programming fibroblasts into iBlastoids’, Nature, vol. 591, no. 7851, pp. 627–632, doi:10.1038/s41586-021-03372-y
  • Xiaodong Liu, John F. Ouyang, Fernando J. Rossello, Jia Ping Tan & Kathryn C. Davidson et al. 2020, ‘Reprogramming roadmap reveals route to human induced trophoblast stem cells’, Nature, vol.586, no.7827, pp.101–107, doi:10.1038/s41586-020-2734-6
  • Polo, Jose M., Susanna Liu, Figueroa, Maria Eugenia, Warakorn Kulalert & Sarah Eminli et al. 2010, ‘Cell type of origin influences the molecular and functional properties of mouse induced pluripotent stem cells’, Nature Biotechnology, vol. 28, no. 8, pp. 848–855,doi:10.1038/nbt.1667
  • Sam Buckberry, Xiaodong Liu, Daniel Poppe, Jia Ping Tan & Guizhi Sun et al. 2023, ‘Transient naïve reprogramming corrects hiPS cells functionally and epigenetically’, Nature, vol.620, no.7975, pp.863–872, doi:10.1038/s41586-023-06424-7

Student projects

The Polo lab has multiple projects spanning development and cancer with both wet lab and computational components. Please contact Professor Polo for more information and further discussions on the projects below:

  • Understanding the epigenetic and transcriptional changes during reprogramming of cells to pluripotent stem cells

    Project supervisor(s): Prof Jose Polo
    Suitable for: Honours, MPhil, PhD
    Location of project: Adelaide Health & Medical Sciences Building, North Terrace, Adelaide

    Description:
    The derivation of human embryonic stem cells (hESCs) and, more remarkably, the generation of human induced pluripotent stem cells (iPSCs) has revolutionised our understanding of pluripotency and opened new avenues for disease modelling, drug screening and regenerative medicine. The ability to reprogram any mature cell back to a pluripotent state, and then into another cell type, provides a unique opportunity to dissect the molecular and cellular events that occur during this conversion. Our lab aims to understand the kinetics and universality of the epigenetic and genomic changes occurring during reprogramming, the composition and assembly kinetics of transcriptional regulation complexes of pluripotency genes and how the cell of origin influences the in vitro and in vivo plasticity potential of cells generated during the reprogramming process.

    We will achieve this by combining stem cell technologies with several different molecular, biochemical, cellular techniques and genome-wide approaches; including ChIP, CUT&RUN, and single cell “omics”.

  • Modelling early placentation

    Project supervisor(s): Prof Jose Polo
    Suitable for: Honours, MPhil, PhD
    Location of project: Adelaide Health & Medical Sciences Building, North Terrace, Adelaide

    Description:
    The derivation of human embryonic stem cells (hESCs) and, more remarkably, the generation of human induced pluripotent stem cells (iPSCs) has revolutionised our understanding of pluripotency and opened new avenues for disease modelling, drug screening and regenerative medicine.

    However, the trophectoderm (TE) gives rise to the placenta and as such, iPSCs cannot provide models for TE or placenta. To address this important challenge, our lab has generated iTSCs for the first time. Our lab will use this model with both human and non-human primate cells, to understand the transcriptional and epigenomic changes that occur in vitro in the early human trophoblast and demonstrate that iTSCs can be used for disease modelling.

  • Pluripotency factors and cancer

    Project supervisor(s): Prof Jose Polo
    Suitable for: Honours, MPhil, PhD
    Location of project: Adelaide Health & Medical Sciences Building, North Terrace, Adelaide

    Description:
    Pluripotent stem cells can self-renew indefinitely and give rise to all cells of the adult organism.  These remarkable capacities are the result of a core transcriptional network controlled by OCT4, SOX2 and NANOG, whose expression is lost upon differentiation. Several cancers have been shown to reactivate OCT4, SOX2 and/or NANOG, with high expression levels positively correlating with cancer progression and severity.  Since they are linked to proliferative and multi-lineage differentiation capacity of cancer cells, they present attractive anticancer targets.  However, they are considered “undruggable” since they lack catalytic active sites for molecules to bind.

    To provide therapeutic alternatives, the Polo research group has adapted and developed various novel techniques to determine how expression and function of the pluripotency factors OCT4, SOX2 and NANOG are controlled in various physiological and pathological cell types, including embryonic stem cells and cancer respectively.

    The candidate will develop skills in a combination of different molecular, biochemical and cellular techniques and use genome wide approaches (RNA-seq, MS-MS, ATAC-seq, ChIP-seq, SC-RNA-seq, etc.) to dissect the nature and dynamics of such events.

    • Uncovering the regulatory complex of Bcl6 in lymphomas

      Project supervisor(s): Prof Jose Polo
      Suitable for: Honours, MPhil, PhD
      Location of project: Adelaide Health & Medical Sciences Building, North Terrace, Adelaide

      Description:
      Cellular identity is controlled by transcription factors (TFs), which bind to specific regulatory elements (REs) within the genome to regulate gene expression and cell fate changes.  Recent advances in epigenome profiling techniques have significantly increased our understanding of which REs are utilised in which cell type, however, which factors interact with these REs remains largely elusive.

      A major impediment to dissecting protein complexes at specific genomic loci is the shortage of appropriate techniques.  The most common technique to assess TF binding is chromatin immunoprecipitation (ChIP), which relies on antibodies to interrogate the binding sites of a single TF.  Yet, ChIP does not allow dissection of the composition of a multi-protein complex at a specific locus.

        Graphical representation of TINC: TALE-mediated isolation of nuclear chromatin

        Graphical representation of TINC: TALE-mediated isolation of nuclear chromatin

        Importantly, the Polo research group has developed a novel epigenetic technique termed TINC (TALE-mediated Isolation of Native Chromatin), which allows us to do exactly that (Knaupp et al., Stem Cell Reports 2020).  TINC relies on epitope-tagged TALEs, which are DNA-binding proteins engineerable to target specific genomic regions. Upon cross-linking of the cells, the target regions are isolated based on affinity purification of the TALE and associated nucleic acid and protein molecules are analysed by next generation sequencing and mass spectrometry, respectively.

        In our proof-of-concept experiments, we dissected the protein complex formed at the Nanog promoter, a key pluripotency RE.  We identified TFs previously known to bind to this locus as well as novel proteins whose role in pluripotency we further validated (Knaupp et al., Stem Cell Reports 2020).  Consequently, with this valuable technique at hand, this PhD project aims at deciphering how the BTB/POZ transcriptional repressor and oncogene BCL6 is (mis)regulated in B-cell lymphomas, which in turn has major potential in identifying novel therapeutic targets.

        The candidate will develop skills in a combination of different molecular, biochemical and cellular techniques and genome wide approaches (RNA-seq, MS-MS, ATAC-seq, ChIP-seq, SC-RNA-seq, etc.) to dissect the nature and dynamics of such events.

        This project has both wet laboratory and bioinformatic components.