Mike Clark from The University of Melbourne in Australia presented at London Calling 2024 on “Igniting single-cell analysis of full-length RNA isoforms in brain development.” Clark works on RNA isoforms and developing single-cell analysis methods. They noted that almost all human genes express multiple RNA isoforms. Clark emphasized that alternative splicing modifies which exons are included in the processed mRNA. The Clark research group works on molecular mechanisms controlling brain development and what happens in neurodevelopmental and neuropsychiatric disorders. The team wants to identify isoforms that may be crucial for brain development. Clark explained that bulk RNA sequencing is an average of the cells in a sample. Single-cell transcriptomics provides the resolution to learn about the expression profiles of different cell types. Clark and the team wanted to develop a long-read scRNA-seq method specific for isoform profiling. The lab developed software and methods. The Flames software package is used with 10X Genomics single-cell platforms and has been undergoing updates. Blaze is additional software the team developed. Clark used a neuron development model and applied their methods and analyses with Flames and Blaze. Samples were sequenced on PromethION flow cells to obtain a depth of 30,000 reads per cell. The data allowed for the identification of cell types and the development trajectory. The early samples are stem cells and progress to neuron types. Clark ended the session by discussing novel isoforms they identified and isoform-switching events during brain development. For example, the PKM novel isoform they found is expressed only in neurons, while other isoforms of PKM are found in progenitors. The tools the Clark lab developed can detect cell types and developmental progression. Isoform dynamics are now detectable on a single-cell level!
