Neural crest migration is driven by a few trailblazer cells with a unique molecular signature narrowly confined to the invasive front

Abstract

Neural crest (NC) cell migration is crucial to the formation of peripheral tissues during vertebrate development. However, how NC cells respond to different microenvironments to maintain persistence of direction and cohesion in multicellular streams remains unclear. To address this, we profiled eight subregions of a typical cranial NC cell migratory stream. Hierarchical clustering showed significant differences in the expression profiles of the lead three subregions compared with newly emerged cells. Multiplexed imaging of mRNA expression using fluorescent hybridization chain reaction (HCR) quantitatively confirmed the expression profiles of lead cells. Computational modeling predicted that a small fraction of lead cells that detect directional information is optimal for successful stream migration. Single-cell profiling then revealed a unique molecular signature that is consistent and stable over time in a subset of lead cells within the most advanced portion of the migratory front, which we term trailblazers. Model simulations that forced a lead cell behavior in the trailing subpopulation predicted cell bunching near the migratory domain entrance. Misexpression of the trailblazer molecular signature by perturbation of two upstream transcription factors agreed with the in silico prediction and showed alterations to NC cell migration distance and stream shape. These data are the first to characterize the molecular diversity within an NC cell migratory stream and offer insights into how molecular patterns are transduced into cell behaviors.

Resolving in vivo gene expression during collective cell migration using an integrated RNAscope, immunohistochemistry and tissue clearing method

“…Second, using the example of SOX10, ITGB5, and NBL1 we quantified their regional expression differences within a typical cranial neural crest cell migratory stream and within single cells. High expression of ITGB5 in lead neural crest cells confirmed our RT-qPCR results (McLennan et al, 2015) and provided confidence that ITGB5 signaling may be important for neural crest cell invasion. Third, spot detection of SOX10, ITGB5, and NBL1 transcripts in HNK1-positive neural crest cells showed most cells express all three genes and detection of spots is not solely dependent on the cell size.…”

“…Furthermore, our single cell RT-qPCR analysis revealed a set of 16 out of 96 genes highly expressed and consistent during migration in cells restricted to the invasive front (McLennan et al, 2015). This heterogeneity of expression is driven by microenvironmental signals as cells travel through different microenvironments (McLennan et al, 2015). In order to better understand the molecular underpinnings of this neural crest cell invasion signature and gene expression heterogeneity within a typical migratory stream, it was clear we needed to develop a rapid means to visualize multiple genes within individual migrating neural crest cells in the same embryo as cells traveled to deep peripheral locations.…”

“…We previously discovered by RT-qPCR that migrating cranial neural crest cells have a robust spatial heterogeneity of gene expression that depends on cell position within a typical migratory stream (McLennan et al, 2015). Furthermore, our single cell RT-qPCR analysis revealed a set of 16 out of 96 genes highly expressed and consistent during migration in cells restricted to the invasive front (McLennan et al, 2015). This heterogeneity of expression is driven by microenvironmental signals as cells travel through different microenvironments (McLennan et al, 2015).…”

Mapping a multiplexed zoo of mRNA expression

“…The fact that amplification polymers carry up to hundreds of fluorophores (Choi et al, 2014) makes it possible to achieve high signal-to-background even when autofluorescence is high [e.g. in whole-mount vertebrate embryos (Choi et al, 2014;McLennan et al, 2015;Huss et al, 2015), in thick mouse brain sections (Sylwestrak et al, 2016) or in bacteria contained within environmental samples or other organisms (Rosenthal et al, 2013;Yamaguchi et al, 2015;Nikolakakis et al, 2015)]. The resulting HCR signal is stable for at least 1 week in zebrafish embryos stored in solution (Fig.…”

Neural crest and cancer: Divergent travelers on similar paths

“…More recently, a unique molecular signature was found to be consistently expressed throughout migration in a subset of NCCs at the leading edge of the migrating stream. When this molecular signature is perturbed in cranial NCCs, migration distances and patterns are disrupted (McLennan et al, 2015). However, subsequent work has shown that physical ablation of NCCs at the leading edge does not appear to impact cranial NC migration (Richardson et al, 2016).…”