Zebrafish Numb and Numblike Are Involved in Primitive Erythrocyte Differentiation

BACKGROUND:Notch signaling is an evolutionarily conserved regulatory circuitry implicated in cell fate determination in various developmental processes including hematopoietic stem cell self-renewal and differentiation of blood lineages. Known endogenous inhibitors of Notch activity are Numb-Nb and Numblike-Nbl, which play partially redundant functions in specifying and maintaining neuronal differentiation. Nb and Nbl are expressed in most tissues including embryonic and adult hematopoietic tissues in mice and humans, suggesting possible roles for these proteins in hematopoiesis. METHODOLOGY AND PRINCIPAL FINDINGS:We employed zebrafish to investigate the possible functional role of Numb and Numblike during hematopoiesis, as this system allows a detailed analysis even in embryos with severe defects that would be lethal in other organisms. Here we describe that nb/nbl knockdown results in severe reduction or absence of embryonic erythrocytes in zebrafish. Interestingly, nb/nbl knocked-down embryos present severe downregulation of the erythroid transcription factor gata1. This results in erythroblasts which fail to mature and undergo apoptosis. Our results indicate that Notch activity is increased in embryos injected with nb/nbl morpholino, and we show that inhibition of Notch activation can partially rescue the hematopoietic phenotype. CONCLUSIONS AND SIGNIFICANCE:Our results provide the first in vivo evidence of an involvement of Numb and Numblike in zebrafish erythroid differentiation during primitive hematopoiesis. Furthermore, we found that, at least in part, the nb/nbl morphant phenotype is due to enhanced Notch activation within hematopoietic districts, which in turn results in primitive erythroid differentiation defects

In vivo cell biology using Gal4-mediated multicolor subcellular labeling in zebrafish

The behavior of a cell is determined by the interplay of its subcellular components. Thus, being able to simultaneously visualize several organelles inside cells within the natural context of a living organism could greatly enhance our understanding of developmental processes. We have established a Gal4-based system for the simultaneous and cell type specific expression of multiple subcellular labels in transparent zebrafish embryos. This system offers the opportunity to follow intracellular developmental processes in a live vertebrate organism using confocal fluorescence time-lapse microscopy. Using this approach we recently showed that the centrosome neither persistently leads migration nor determines the site of axonogenesis in migrating neurons in the zebrafish cerebellum in vivo. Here we present additional in vivo findings about the centrosomal and microtubule dynamics of neuroepithelial cells during mitotic cleavages at early neural tube stages

Neurons derive from the more apical daughter in asymmetric divisions in the zebrafish neural tube

International audienceIn the developing CNS asymmetric cell division is critical to maintain the balanced production of differentiating neurons while also renewing the population of neural progenitors. In invertebrates this process depends on asymmetric inheritance of fate determinants during progenitor divisions. A similar mechanism is widely believed to underlie asymmetrically fated divisions in vertebrates but compelling evidence for this is missing. We use live imaging of individual progenitors in the intact zebrafish embryo CNS to test this hypothesis. We provide the first direct evidence that asymmetric inheritance of a subcellular domain is strongly correlated with asymmetric daughter fates and reveal an unexpected feature of this process. The daughter cell destined to become a neuron is derived from the more apical of the two daughters, while the more basal daughter inherits the basal process and replenishes the apical progenitor pool