Generation of a new six1-null line in Xenopus tropicalis for study of development and congenital disease

The vertebrate Six (Sine oculis homeobox) family of homeodomain transcription factors play critical roles in the development of several organs. Six1 plays a central role in cranial placode development, including the precursor tissues of the inner ear, as well as other cranial sensory organs and the kidney. In humans, mutations in SIX1 underlie some cases of branchio-oto-renal syndrome (BOR), which is characterized by moderate to severe hearing loss. We utilized CRISPR/Cas9 technology to establish a six1 mutant line in Xenopus tropicalis that is available to the research community. We demonstrate that at larval stages, the six1-null animals show severe disruptions in gene expression of putative Six1 target genes in the otic vesicle, cranial ganglia, branchial arch and neural tube. At tadpole stages, six1-null animals display dysmorphic Meckel's, ceratohyal and otic capsule cartilage morphology. This mutant line will be of value for the study of the development of several organs as well as congenital syndromes that involve these tissues.

Sox8 is sufficient to reprogram ectoderm into ear vesicles and associated neurons

The vertebrate inner ear arises from a pool of progenitors with the potential to give rise to all the sense organs and cranial ganglia of the head1-6. Here we explore the molecular mechanisms that control ear specification from these precursors. Using a multi-omics approach combined with loss-of-function experiments we identify a core transcriptional circuit that imparts ear identity, along with non-coding elements that integrate this information. This analysis places the transcription factor Sox8 at the top of the ear determination network. Introducing Sox8 into cranial ectoderm not only converts non-ear cells into ear progenitors, but also activates the cellular programmes for ear morphogenesis and neurogenesis. Thus, Sox8 emerges as a master regulator of ear identity and may be a key factor for sense organ cell reprogramming.

Expression of the neurotrophic tyrosine kinase receptors, ntrk1 and ntrk2a, precedes expression of other ntrk genes in embryonic zebrafish

Background The neurotrophic tyrosine kinase receptor (Ntrk) gene family plays a critical role in the survival of somatosensory neurons. Most vertebrates have three Ntrk genes each of which encode a Trk receptor: TrkA, TrkB, or TrkC. The function of the Trk receptors is modulated by the p75 neurotrophin receptors (NTRs). Five ntrk genes and one p75 NTR gene (ngfrb) have been discovered in zebrafish. To date, the expression of these genes in the initial stages of neuron specification have not been investigated. Purpose The present work used whole mount in situ hybridization to analyze expression of the five ntrk genes and ngfrb in zebrafish at a timepoint when the first sensory neurons of the zebrafish body are being established (16.5 hpf). Because expression of multiple genes were not found at this time point, we also checked expression at 24 hpf to ensure the functionality of our six probes. Results At 16.5 hpf, we found tissue specific expression of ntrk1 in cranial ganglia, and tissue specific expression of ntrk2a in cranial ganglia and in the spinal cord. Other genes analyzed at 16.5 hpf were either diffuse or not detected. At 24 hpf, we found expression of both ntrk1 and ntrk2a in the spinal cord as well as in multiple cranial ganglia, and we identified ngfrb expression in cranial ganglia at 24 hpf. ntrk2b, ntrk3a and ntrk3b were detected in the developing brain at 24 hpf. Conclusion These data are the first to demonstrate that ntrk1 and ntrk2a are the initial neurotrophic tyrosine kinase receptors expressed in sensory neurons during the development of the zebrafish body, and the first to establish expression patterns of ngfrb during early zebrafish development. Our data indicate co-expression of ntrk1, ntrk2a and ngfrb, and we speculate that these overlapping patterns indicate relatedness of function.

Hmx gene conservation identifies the evolutionary origin of vertebrate cranial ganglia

The evolutionary origin of vertebrates included innovations in sensory processing associated with the acquisition of a predatory lifestyle1. Vertebrates perceive external stimuli through sensory systems serviced by cranial sensory ganglia (CSG) which develop from cranial placodes; however understanding the evolutionary origin of placodes and CSGs is hampered by the gulf between living lineages and difficulty in assigning homology between cell types and structures. Here we use the Hmx gene family to address this question. We show Hmx is a constitutive component of vertebrate CSG development and that Hmx in the tunicate Ciona is able to drive the differentiation program of Bipolar Tail Neurons (BTNs), cells previously thought neural crest homologs2,3. Using Ciona and lamprey transgenesis we demonstrate that a unique, tandemly duplicated enhancer pair regulated Hmx in the stem-vertebrate lineage. Strikingly, we also show robust vertebrate Hmx enhancer function in Ciona, demonstrating that deep conservation of the upstream regulatory network spans the evolutionary origin of vertebrates. These experiments demonstrate regulatory and functional conservation between Ciona and vertebrate Hmx, and confirm BTNs as CSG homologs. Our analysis also identifies derived evolutionary changes, including a genetic basis for secondary simplicity in Ciona and unique regulatory complexity in vertebrates.