The challenge of defining rare genetic programs by single-cell RNA sequencing: Insights from phloem studies

Single-cell RNA sequencing technology provides unprecedented opportunities to capture the spatiotemporal aspects of genetic programs that drive cell differentiation. Phloem is a subtype of vascular tissue that is crucial for transporting sugars, proteins, and other organic molecules. Its critical role in the redistribution of carbon between photosynthetically active "source" organs (leaves, shoots) and "sink" tissues/organs incapable of fixing carbon (wood, storage organs, root system, etc.) has made phloem development and its transcriptional regulation a prime research focus. Developing phloem cells are strongly underrepresented in whole-organ samples. In the root tip, the functional phloem unit consists of only a single cell file of sap-conducting sieve elements surrounded by metabolically active CCs and phloem pole pericycle cells, which assist in sap unloading. These cell types display significant transcriptional similarities to each other early in their developmental trajectories, which makes separation of the corresponding transcriptomic data very challenging. Moreover, the most distinct process of phloem sieve element differentiation, the removal of the cell nucleus, is completed within a few hours, making it an extremely rare event. Recent advances in single-cell RNA sequencing (scRNA-seq) inaugurated the process of high-resolution characterization of developmental processes of various tissues. We believe the insights from the analysis of phloem could be informative for analyzing the development of other cell types that have only minor representation in large single-cell transcriptomics datasets.