@phdthesis{discovery1324536,
            note = {Unpublished},
           title = {Genetic studies of inner ear development in Xenopus tropicalis},
          school = {UCL (University College London)},
           month = {August},
            year = {2011},
          author = {Ironfield, H. V.},
             url = {https://discovery.ucl.ac.uk/id/eprint/1324536/},
        abstract = {Xenopus tropicalis provides an excellent model system for studying inner ear
development. At ear-forming stages the head is transparent, so this organ can
easily be observed, and the small genome, short generation time, and
genomic resources makes this species amenable to genetic studies. A group
of recessive mutants displaying inner ear defects were obtained from a pilot
mutagenesis and gynogenetic screen. The primary inner ear defect was
identified for each mutant and each lesion mapped to a chromosome arm.
Two mutants, seasick (ssk) and komimi (kom) were selected for high
resolution positional cloning and phenotypic characterization.
The ssk mutant was identified due to enlarged otoconial crystals, decreased
pigmentation, and balance defects. Positional cloning showed that the ssk
phenotype is caused by a premature stop codon in the {\ensuremath{\delta}} subunit of the
adapter protein (AP)-3 complex. AP-3 is required to transport proteins from the
endosome and golgi to lysosome-related organelles, a class of organelles that
degrade intracellular proteins but also have cell-specific functions such as
ionic regulation and pigment synthesis. ap3{\ensuremath{\delta}}1 is expressed in the
endolymphatic sac, which regulates the ionic composition of the fluid filling the
inner ear, suggesting that the otoconial defect may be the result of an ionic
imbalance. AP-1 also transports proteins from the endosome to the lysosomerelated
organelles. Upregulation of AP-1 subunits are observed in response to
ssk, suggesting that AP-1 may compensate for loss of or decreased levels of
AP-3 function.
In kom mutants a single large otoconium forms over each macula, and
tadpoles exhibit strong balance defects such as circular swimming. Also, the
cells comprising the macula are dysmorphic and lack the columnar structure
found in wild type. Positional cloning shows that the kom phenotype is caused by a splicing defect causing partial loss of the otoconin-90 signal peptide and
likely stops incorporation of the protein into the core of otoconial crystals.
This work successfully demonstrates that X. tropicalis forward genetics is a
valuable tool for studying the development of the inner ear.}
}