Figure 1. The schematic diagram represents how histone modifications, DNA methylation, non-coding RNAs and regulatory elements maintain 3D genome organization and regulate gene expression. (Courtesy- PPD lab).
Epigenetic modifications (DNA and histone modifications) are highly dynamic and regulate gene expression patterns during maintenance of cellular state, differentiation and development and in response to environmental signals. Epigenetic modifications are constantly reshaping the chromatin architecture, 3D structure and accessibility of genes to the transcription machinery for gene expression (Figure 1). We are interested in exploring how gene expression programs are regulated in order to control embryonic stem cell (ESC) state, differentiation and development; and how deregulation of gene expression programs cause diseases.
Ongoing and future projects in our lab include:
1. Dissecting the combinatorial functions of histone demethylases in transcriptional regulatory networks in mouse ESCs (mESCs)
2. Identifying novel epigenetic regulators using CRISPR-Cas9 mediated functional genetic screen and investigating their precise functions in mESCs
3. Examining the detailed functions of Enhancer and Super-enhancer regulatory elements in mESCs
4. Understanding the roles of 3D genome organization in transcription regulation
5. Investigating the epigenetic dynamics in normally developing brain and in neurodevelopmental disorders using the mouse and human brain organoids
RNA epigenetics or Epitranscriptomics:
Eukaryotic RNAs carry more than 150 distinct chemical modifications, some having been demonstrated as being reversible. The understanding of each of these RNA modifications is incomplete and just beginning to emerge. The majority of the RNA modifications have been characterised in non-coding RNAs, including transfer RNAs, ribosomal RNAs, small nuclear RNAs and, to some extent, messenger RNAs. Advancement of next-generation sequencing tools has empowered the identification of several distinct internal RNA modifications on a transcriptome-wide scale, including 6-methyladenosine (m6A), 1-methyladenosine, pseudouridine (Ψ) and 5-methylcytosine (m5C)— these are the four most commonly occurring and extensively investigated RNA modifications to date (Figure 2). Recent studies have begun to elucidate the molecular and biological functions of these internal RNA modifications in many organisms, from yeast to mammals. Several RNA modifications have already been linked to human diseases, including cancer, cardiovascular disease, neurological disorders, metabolic disease, mitochondrial-related defects and genetic birth defects. We are interested in learning how these RNA modifications can regulate genetic information at the transcriptional, post-transcriptional and translational levels in the context of ESC state maintenance, development and diseases.
Ongoing and future projects in our lab include:
1. Investigating the role of RNA modifications in ESCs and other differentiated cell types
2. Identifying new factors (Writes, Erasers, and Readers) in m6A pathways and studying their detailed mechanism of action in m6A RNA modification and gene regulation in ESCs
3. Dissecting the role of RNA modifications in normally developing brain and in neurodevelopmental disorders using brain organoids
4. Understanding the role of RNA modifications in cancer
Figure 2. Most abundant internal chemical modifications in mRNAs; and their distribution. (Courtesy- PPD lab).
3D brain organoids to investigate normal neurodevelopment and neurodevelopmental disorders (NDDs):
Three-dimensional (3D) brain organoids derived from embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs), appear to resemble many aspects of spatial organization and functionality of the developing brain. Brain organoids or cerebral organoids reflecting the broad regional identities. Alternatively, they may contain structures that resemble specific brain regions, therefore can be referred to as organoids of that regions– such as forebrain organoids, midbrain organoids etc. Recent advancement of the brain organoid generation from ESCs/iPSCs (or patient-derived iPSCs) provide a new tool to model both “normal” and “pathological” human brains, and that eventually enhance our ability to study human brain biology and diseases. Currently, we are using brain organoids to study the roles of epigenetics and epitranscriptomics in normal neurodevelopment and neurodevelopmental disorders (NDDs), as well as in neurological disease modelling. Besides, we are also using CRISPR-Cas9 genome editing tools to manipulate genes/genome in ESCs and iPSCs (or patient-derived iPSCs) to check their effect in the brain organoids. This will be immensely important for studying neurodevelopment, NDDs, disease modelling and personalized medicine.
Figure 3. Immunostaining of an entire mouse cortical organoid represent cortical markers Pax6 (neural progenitors), Tuj1 (neurons) on day 10 (A). Pax6+ (neural progenitors), Tuj1+ (neurons) rosettes on day 10 (B). (Unpublished, PPD lab).
Figure 4.. Immunostaining reveal an entire human cerebral organoid that expresses neural progenitors (Pax6, green) and neurons (Tuj1, red); DAPI, blue, on day 40 (A). Immunostaining of the cortical markers Pax6 (neural progenitors), Tbr2 (migrating neurons), Tbr1 and Ctip2 ( mature neurons) on day 40 (B). (Unpublished, PPD lab).
Figure 5. 3D imaging of Pax6+ (neural progenitors, green), Tuj1+ (neurons, red) rosettes from human cerebral organoid on day 40 (B). (Unpublished, PPD lab).