Rhombomere boundaries are Wnt signaling centers that regulate metameric patterning in the zebrafish hindbrain

Abstract

The vertebrate hindbrain develops from a series of segments (rhombomeres) distributed along the anteroposterior axis. We are studying the roles of Wnt and Delta-Notch signaling in maintaining rhombomere boundaries as organizing centers in the zebrafish hindbrain. Several wnt genes (wnt1, wnt3a, wnt8b, and wnt10b) show elevated expression at rhombomere boundaries, whereas several delta genes (dlA, dlB, and dlD) are expressed in transverse stripes flanking rhombomere boundaries. Partial disruption of Wnt signaling by knockdown of multiple wnt genes, or the Wnt mediator tcf3b, ablates boundaries and associated cell types. Expression of dlA is chaotic, and cell types associated with rhombomere centers are disorganized. Similar patterning defects are observed in segmentation mutants spiel-ohne-grenzen (spg) and valentino (val), which fail to form rhombomere boundaries due to faulty interactions between adjacent rhombomeres. Stripes of wnt expression are variably disrupted, with corresponding disturbances in metameric patterning. Mutations in dlA or mind bomb (mib) disrupt Delta-Notch signaling and cause a wide range of patterning defects in the hindbrain. Stripes of wnt1 are initially normal but subsequently dissipate, and metameric patterning becomes increasingly disorganized. Driving wnt1 expression using a heat-shock construct partially rescues metameric patterning in mib mutants. Thus, rhombomere boundaries act as Wnt signaling centers required for precise metameric patterning, and Delta signals from flanking cells provide feedback to maintain wnt expression at boundaries. Similar feedback mechanisms operate in the Drosophila wing disc and vertebrate limb bud, suggesting coaptation of a conserved signaling module that spatially organizes cells in complex organ systems.

Frizzled-3a and Wnt-8b genetically interact during forebrain commissural formation in embryonic zebrafish

“…To test this idea we knocked down zebrafish Wnt8b using previously described antisense translational-blocking morpholinos (MOs) (Riley et al, 2004). While injection of 2ng of wnt8b MO resulted in embryos with abnormal gross morphology (data not shown), we found most embryos developed normally following injection of 0.5-1.0ng of wnt8b MO.…”

“…Standard control (Std-cont MO) (5'-CCT CTT ACC TCA GTT ACA ATT TAT A-3) was designed and synthesized by Gene tools. Previously described MOs against fzd3a -MO (5'-AGA ACC ATG GTG CGT CCG TCT GTG C-3'; ) and wnt8b-MO (5'-AGG GAG ACT TTC TTC ACC TTT CAC-3'; (Riley et al, 2004)) were used.…”

Expression of protocadherin 18 in the CNS and pharyngeal arches of zebrafish embryos

“…3), similar to the radial glia (Trevarrow et al, 1990). Some signaling and transcription factors such as deltaA (Delta-Notch signaling), rasgef1b (Ras GEF), and beta3.1 (bHLH), have been reported to show similar expression patterns in the hindbrain (Adolf et al, 2004, Epting et al, 2007, Riley et al, 2004. Although functional relationships between these signals and pchd18 remain to be elucidated, it is interesting to assume that pcdh18 is involved in rhombomeric boundary formation driven by Delta-Notch signaling.…”

Fishing for gene function – endocrine modelling in the zebrafish

“…This is now being made possible by the availability of heat-shock-inducible vectors that can be injected into the embryo; however, a potential problem may lie in the fact that such vectors show a mosaic distribution in the embryo, and thus ubiquitous and uniform overexpression may be difficult to achieve (Riley et al 2004). Currently, inducible gene expression systems such as this are not widely used in the adult animal, and that is a distinct disadvantage to many endocrinologists.…”