Hmx4 regulates Sonic hedgehog signaling through control of retinoic acid synthesis during forebrain patterning

Mutations in H6-homeobox (HMX) genes are linked to neural mispatterning and neural tube closure defects in humans. We demonstrate that zebrafish Hmx4 regulates the signaling of two morphogens critical for neural development, retinoic acid (RA) and Sonic hedgehog (Shh). Hmx4-depleted embryos have a strongly narrowed eye field and reduced forebrain Shh target gene expression. hmx4 morphants fail to properly transcribe the Shh signal transducer gli3, and have reduced ventral forebrain specification. Hmx4-depleted embryos also have neural tube patterning defects that phenocopy RA-deficiency. We show that Hmx4 is required for the initiation and maintenance of aldh1a2, the principal RA-synthesizing gene. Loss of RA is the primary defect in Hmx4-depleted embryos, as RA treatment rescues a number of the neural patterning defects. Surprisingly, RA treatment also rescues forebrain morphology, gli3 transcription, and Shh signaling. We propose that Hmx4 is a critical regulator of retinoic acid synthesis in a developing embryo, and that this regulation is essential for controlling Shh signaling and forebrain development.

Research highlights
► We performed the first loss-of-function analysis of the H6 homeobox gene hmx4.
► hmx4 zebrafish morphants have a narrowed forebrain, and reduced Shh and RA signaling.
► Hmx4 regulates RA synthesis.
► RA treatment rescues both the RA- and Shh-deficient phenotypes.
► Thus, Hmx4-regulated RA synthesis is essential for Shh-dependent forebrain patterning.