Deciphering Disease Mechanisms through Functional Genomics

-- Dr. Vijay Tiwari --

The Brain- a three-pound of flesh that resides in the skull- is undoubtedly the most complex thing of all objects in the universe. This complexity poses challenges to study the underlying mechanisms responsible for causing brain disorders. If applied well, the functional genomics toolbox can help in discerning the workings of this mysterious organ along with deciphering the genetic basis of brain disorders. Prof. Vijay Tiwari, Professor at the Queen’s University (UK) and Southern University of Denmark (Denmark) gave a talk entitled “ Deciphering Disease Mechanisms through Functional Genomics” to discuss how his research lab leveraged functional genomics to study brain disorders. The talk was jointly organized by IBSE and RBCDSAI on 21st June 2022.

Prof. Tiwari commenced his talk by describing the structure of the brain and told the audience that, unlike the rat’s brain which is small and flat, the human brain is large, folded and has a neocortex which is a newly evolved region of the brain which has expanded over the course of evolution to give us intelligence. He next mentioned that telencephalon which has around 80 cell types gives rise to the majority of the neocortex and posed questions such as what mechanisms govern this process? how do these processes go wrong in diseases? Why do we need to know the developmental mechanisms? He then answered the question by saying that when such mechanisms malfunction then it leads to diseases such as autism, fetal alcohol spectrum disorders, mental retardation, Asperger’s syndrome etc and the knowledge of mechanisms is also required for improved therapeutic strategies- for neurological disorders or for regenerative therapy via re-programming.

Next, he talked about the neuro-differentiation mechanism. Comparing, the mouse brain with the human brain, he said that the human sub-ventricular zone has two other cell types and it has expanded into the inner subventricular zone and outer sub-ventricular zone. Elaborating on the differentiation pathway, he said that we have apical progenitors which differentiate into basal progenitors which further differentiate to give rise to neurons that settle in the cortical regions and make synapsis and connections. Further, elaborating on the differentiation mechanism, he said that the transition from progenitor to cell i.e. changes in cell fate requires re-programming of gene expression that involves epigenetic mechanisms such as DNA methylation, sumolyation, regulatory non-coding RNA etc.

He next explained how the epigenetic changes lead to the expression of genes in a cell. He told that initially in an undifferentiated neural progenitor cell, histones are compact and no gene expression takes place. However, when the trigger for differentiation occurs, the trigger targets a factor X to the loci which bring epigenetic machinery to change the repressive histone modification to active modification. With this histone modification, chromatin now becomes accessible due to displacement of histones and is available for binding of transcription factors. These transcription factors allow for binding of basal machinery such as polymerases for gene expression and this expression of differentiation-specific genes then orchestrates the steps in the transition of a progenitor cell to neuron.

Further, he mentioned that his lab works on various levels of differentiation pathway i.e. differentiation signals, chromatin accessibility and gene expression to study brain development. He next mentioned his research work on active and primed enhancers and said that his team performed whole-genome bisulfite sequencing- a method that looks at the DNA methylation at a single base-pair resolution. Their research found that majority of the genome is methylated which corroborates with the fact that most of the genome is in a heterochromatin state. However, certain DNA regions showed a drop in methylation and these regions they found were enhancers. The team also found a cluster of enhancers that were in the vicinity of each other which they called super enhancers and showed that when neurons are activated in the brain they re-model super enhancers to activate a different set of genes. The team also showed that during development processes such as neurogenesis, a factor called NeuroD1 is transiently induced which targets distal regions and activate enhancers genome-wide leading to the expression of many genes. He further told that the team next sought out to find that why only certain regions in the genome are selected to be enhancers and decided to study if the DNA region containing enhancers has a propeller twist. His team carried out an experiment which showed that propeller twist highly correlates with the DNA surface accessibility. Also, they did atomic force microscopy on enhancer DNA region containing propeller twist and control DNA and found that the propeller twist regions have more bendability and high propeller twist containing regions are more amenable to mutations as they are more accessible. They also found that propeller twist levels correlate with the nuclear organization of epigenetic states and it was found that regions with higher levels of acetylation have more propeller twist and propeller twist is high at the centre of the enhancers. The research group also found that cell fate switches/ reprogramming involves transient usage of propeller twist regulatory elements and found that SNPs in enhancers increase the risk of neurological disorders.

He next talked about his other research project where the team measured active enhancers throughout the human brain and found that SNPs in the enhancer of the PHF21B gene is associated with intellectual disability and depression. Elaborating on the function of this gene, he said that they found when cells transit from progenitor state to differentiated state, this gene binds to cell cycle genes and recruits repressive epigenetic machinery to deactivate these genes which is essential to shut down the cell cycle genes. Therefore, a mutation in this gene causes the cells to continue to proliferate and not differentiate which causes intellectual disability and depression.

Next, he explained his research on the question of why brain folding is essential? He told that to study this problem, they chose Ferrets as an organism as it has cortical folding. The team took regions of gyri and sulcus from the Ferret brain and did whole-genome transcriptomics and lysine 27 acetylation studies. They found that although these two regions in the brain have the same cell types, they have a different transcriptomic profiles as gene regulatory processes play an important part in the formation of folds. Also, the genes that are differentially expressed between gyrus and sulcus include many genes associated with several neurodevelopmental disorders which point to the fact that neuro-disorders can arise due to aberrant cortical folding programs. This observation is in concordance with MRI studies that have shown that there is an aberrant cortical folding in the brains of autism and epilepsy patients.

Prof. Tiwari then talked about his research work on Epithelial-Mesenchymal Transition (EMT) mechanism and said that through the single-cell RNA technique they have found that a number of EMT genes involved in developmental processes are also relevant in cancer. They found that Fbox32 is a gene essential for cell migration during neurogenesis and metastasis and showed through the rat study that if Fbox32 is mutated then cancer does not metastasize. The team also studied EMT genes in the context of chemo-resistant breast cancer and has patented 20 signature genes which will be used to classify if a woman will be chemo-resistance to breast cancer in future.

He next said that his team is also using single-cell techniques to understand common causes of neurodevelopmental disorders. Around 70% of autistic kids have ADHD and the frontal cortex is affected in these kids. They found that genes differentially expressed in Autism Spectrum Disorder co-morbidity groups are linked to neurodevelopmental processes and have shortlisted six novel genes for ASD from that study that show high neuronal expression and interesting developmental trajectory and aim to knock them to get the phenotype.

Prof. Tiwari concluded his lecture by talking about various projects in his lab i.e. identification of transcription factors shaping the epigenetic landscape, characterization of novel epigenetic regulators, dissection of the function of chromatin accessibility in cell fate, single-cell transcriptome/epigenome analysis and spatial transcriptomics, deciphering the disrupted gene regulatory program in neurodevelopmental disorders and system biology of gene regulatory network. The talk was well-received by the audience and a question-answer session followed after the talk.

Keywords

Genomics, Human brain , Epigenetic