Studies of gene expression are typically carried out by a molecular analysis that averages entire populations of cells in culture, or in a tissue. This approach cannot detect cell to cell variability, or collect subcellular information, such as spatial distribution. At the transcriptional level, it is evident that even a robust transcriptional response in ensemble measurements is not uniform among all cells in a population. At the post-transcriptional level, mRNAs and proteins can be trafficked to specific sub-cellular compartments allowing spatiotemporal regulation of gene expression, but these critical spatial relationships are lost with common molecular biology approaches. Through direct visualization of mRNA during the biogenesis process and analyzing the distribution of single mRNA molecules in cells we have gained a deeper understanding of gene expression at many levels. Recent technical advances have made these types of analysis more accessible than ever. The utility of this approach toward studying transcriptional events is underscored throughout many of the articles within this volume. Techniques such as fluorescent in situ hybridization (FISH) are being applied to single molecule studies in fixed cells with far-reaching results, but they are limited in their ability to provide information about the dynamic nature of mRNA in vivo, so methodology to visualize single mRNA molecules in living cells has become desirable. In this article, we will discuss the state-of-the-art tagging systems used for real-time imaging of mRNAs that have been developed. We will present an overview of how these approaches have been applied to impacting our view of gene expression.

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Cell migration requires a complicated network of structural and regulatory proteins. Changes in cellular motility can impact migration as a result of cell-type or developmental stage regulated expression of critical motility genes. Hnrnpab is a conserved RNA-binding protein found as two isoforms produced by alternative splicing. Its expression is enriched in the subventricular zone (SVZ) and the rostral migratory stream within the brain, suggesting possible support of the migration of neural progenitor cells in this region. Here we show that the migration of cells from the SVZ of developing Hnrnpab −/− mouse brains is impaired. An RNA-seq analysis to identify Hnrnpab-dependent cell motility genes led us to Eps8, and in agreement with the change in cell motility, we show that Eps8 is decreased in Hnrnpab −/− SVZ tissue. We scrutinized the motility of Hnrnpab −/− cells and confirmed that the decreases in both cell motility and Eps8 are restored by ectopically coexpressing both alternatively spliced Hnrnpab isoforms, therefore these variants are surprisingly nonredundant for cell motility. Our results support a model where both Hnrnpab isoforms work in concert to regulate Eps8 transcription in the mouse SVZ to promote the normal migration of neural cells during CNS development.