Role of Single Cell mRNA Variation in Systems-Associated Electrically Excitable Cells

The goal of this inter-disciplinary project is to characterize and understand the variability in the expressed transcriptome of human excitable cells. There are two predominant types of excitable cells in the human body, neurons and muscle cells, including heart muscle cells (cardiomyocytes). Many human CNS and heart diseases result from alterations of the electrophysiologic properties of neurons and cardiomyocytes, for example epilepsy and arrhythmia. Although individual neurons and cardiomyocytes exhibit considerable heterogeneity in function, response, and dysfunction, the underlying molecular mechanisms for this heterogeneity are poorly defined. Our preliminary data demonstrate that the variability of the transcriptional profile of single neurons and cardiomyocytes is wide, which is difficult to explain as simple molecular noise. We hypothesize that many transcriptome states correspond to a given cell type, for example a type of neuron or cardiomyocyte. Because a broad set of multi-genic combinations support a particular phenotype, changes in the transcriptome state do not necessarily lead to changes in the phenotype, but may lead to functional heterogeneity between individual neurons and cardiomyocytes. This paradigm explains the variable responses of phenotypically identical cells to application of therapeutic molecules. To test our hypothesis, we will investigate the extent of single cell variation for the whole transcriptome for neurons and cardiomyocytes. We will achieve this in their natural environment using a novel in situ mRNA capture method (TIVA-tag) and functional genomics techniques developed in the Eberwine and Kim labs (TIPeR). This approach will permit an accurate assessment of the extent of multigenic transcriptome variation. We will then systematically perturb the transcriptome of individual neurons and cardiomyocytes and determine the functional effect by examining action potentials.