Fgf3 and Fgf8 dependent and independent transcription factors are required for otic placode specification

Abstract

The vertebrate inner ear develops from the otic placode, an ectodermal thickening that forms adjacent to the presumptive hindbrain. Previous studies have suggested that competent ectodermal cells respond to signals from adjacent tissues to form the placode. Members of the Fgf family of growth factors and the Dlx family of transcription factors have been implicated in this signal-response pathway. We show that compromising Fgf3 and Fgf8 signaling blocks ear development; only a few scattered otic cells form. Removal of dlx3b, dlx4b and sox9a genes together also blocks ear development, although a few residual cells form an otic epithelium. These cells fail to form if sox9b function is also blocked. Combined loss of Fgf signaling and the three transcription factor genes, dlx3b,dlx4b and sox9a, also completely eliminates all indications of otic cells. Expression of sox9a but not dlx3b, dlx4b or sox9b requires Fgf3 and Fgf8. Our results provide evidence for Fgf3-and Fgf8-dependent and -independent genetic pathways for otic specification and support the notion that Fgf3 and Fgf8 function to induce both the otic placode and the epithelial organization of the otic vesicle.

Genetic interactions underlying otic placode induction and formation

“…Recent studies suggest that FGF-mediated signals act as the major inductive component in this process (Phillips et al, 2001;Leger and Brand, 2002;Maroon et al, 2002). In zebrafish, the transcription factors foxi1 and dlx3b/4b are required in the responding ectoderm during the inductive phase (Solomon and Fritz, 2002;Liu et al, 2003;Nissen et al, 2003;Solomon et al, 2003a). The data presented here suggest that foxi1 and dlx3b/4b function in independent pathways and that two early phases of FGF induction are genetically separable.…”

“…Otic Up-Regulation of dlx3b and dlx4b Expression Is Delayed in hsy hsy mutants and dlx3b/4b morpholino-injected embryos (morphants) exhibit a reduction in otic vesicle size, although this defect is generally more severe for dlx3b/4b morphants (Solomon and Fritz, 2002;Liu et al, 2003;Nissen et al, 2003;Solomon et al, 2003a). Both also display a loss of pax2a expression in the otic precursor cells at the three-somite stage (3s), demonstrating an early function for foxi1 and dlx3b/4b in the formation of the otic placode.…”

“…4C) and in dlx3b/4b double morphants ( Fig. 4D) and deletion mutants (Solomon and Fritz, 2002;Liu et al, 2003). However, weak pax2a expression is observed (an embryo from an ace ϩ/-incross displaying a wild-type phenotype; E), ace -/-homozygous mutant (F), ace ϩ/?…”

“…It has become clear that FGF-derived signals constitute a significant part of this inductive capacity (Represa et al, 1991;Ladher et al, 2000;Vendrell et al, 2000;Phillips et al, 2001;Leger and Brand, 2002;Maroon et al, 2002;Liu et al, 2003). In zebrafish, fgf3/8 are expressed in mesendoderm and hindbrain and appear to function as inductive signals at multiple points during otic placode development Phillips et al, 2001;Leger and Brand, 2002;Maroon et al, 2002;Liu et al, 2003). For example, treatment of zebrafish embryos with the FGF inhibitor SU5402 revealed distinct requirements for FGF signaling during early induction and later differentiation (Leger and Brand, 2002).…”

“…Expression becomes progressively restricted and up-regulated in the otic and olfactory primordia during early somitogenesis. Combined loss of function for dlx3b/4b leads to a reduction or absence of otic placodes and a loss of pax2a expression in otic precursor tissue, although pax8 expression initiates normally (Solomon and Fritz, 2002;Liu et al, 2003). Zebrafish foxi1, a Forkhead domain-containing transcription factor (Weigel and Jackle, 1990), is also expressed within otic precursor tissue and appears to function even earlier than dlx3b/4b (Solomon et al, 2003a).…”