Notch signaling regulates midline cell specification and proliferation in zebrafish

Abstract

Notochord and floor plate cells are sources of molecules that pattern tissues near the midline, including the spinal cord. Hypochord cells are also found at the midline of anamniote embryos and are important for aorta development. Delta-Notch signaling regulates midline patterning in the dorsal organizer by inhibiting notochord formation and promoting hypochord and possibly floor plate development, but the precise mechanisms by which this regulation occurs are unknown. We demonstrate here that floor plate and hypochord cells arise from distinct regions of the zebrafish shield. Blocking Notch signaling during gastrulation entirely prevented hypochord specification but only reduced the number of floor plate cells that developed compared to control embryos. In contrast, elevation of Notch signaling at the beginning of gastrulation caused expansion of hypochord at the expense of notochord, but floor plate was not affected. A cell proliferation assay revealed that Notch signaling maintains dividing floor plate progenitors. Together, our results indicate that Notch signaling regulates allocation of appropriate numbers of different midline cells by different mechanisms.

The zebrafish tailbud contains two independent populations of midline progenitor cells that maintain long-term germ layer plasticity and differentiate based on local signaling cues

“…The inhibition of Notch signaling had no effect on floor plate contribution, whereas Notch activation caused a slight increase in the percentage of floor plate cells. These results are consistent with the previously reported gastrula stage role of Notch signaling in promoting floor plate cell proliferation, but not affecting floor plate specification (Latimer and Appel, 2006).…”

“…However, we found that Notch inhibition rescued the enhanced expression of the floor plate marker foxa2 caused by loss of Wnt or overexpression of sox2 (Fig. 5G-L), consistent with the role of Notch in promoting floor plate proliferation (Latimer and Appel, 2006). Additionally, we found that expanded sox2 expression resulting from Wnt inhibition did not depend on Notch function ( Fig.…”

“…To provide more conclusive evidence that endoderm continues to be induced in the tailbud MPCs after gastrulation ends, we tested for continued responsiveness to Notch signaling, which is required for hypochord induction during gastrulation (Appel et al, 1999;Latimer and Appel, 2006;Latimer et al, 2002;Peyrot et al, 2011). We designed heat shock-inducible vectors in the same manner as the β-catenin and sox2 constructs, to allow inducible expression of the Notch intracellular domain (NICD) or a dominant-negative Suppressor of Hairless [dnSu(H)] (Wettstein et al, 1997), allowing temporal cell-autonomous activation or inhibition of the Notch signaling pathway in transplanted cells, respectively.…”

“…There is strong evidence for a conserved role of Notch signaling in regulating hypochord allocation during gastrulation, but the signals regulating medial floor plate allocation are less clear (Appel et al, 1999;Latimer and Appel, 2006;Latimer et al, 2002;Peyrot et al, 2011). Speciesspecific roles for Notch and Midkine signaling have been identified, but a conserved pathway regulating this process across all species has yet to be identified (Gray and Dale, 2010;Lopez et al, 2003Lopez et al, , 2005Peyrot et al, 2011;Schafer et al, 2005).…”

A gene regulatory network directed by zebrafish No tail accounts for its roles in mesoderm formation

“…The Notch pathway is essential for tailbud outgrowth, for differentiating between the hypochord and notochord, and for correct segmentation of the paraxial mesoderm into somites (37,38). dld is a known direct target of Ntl (52), and we also see binding to the upstream regions of dlc, notch3, her1, her11/5, and her12 (data not shown).…”