Otx but Not Mitf Transcription Factors Are Required for Zebrafish Retinal Pigment Epithelium Development

<div><p>Otx and Mitf transcription factors have been implicated in the development of the retinal pigmented epithelium (RPE), but the relationship between these factors and their specific roles in the development of the RPE have not been fully defined. The role of the three Otx transcription factors (Otx1a, Otx1b, and Otx2) and two Mitf transcription factors (Mitfa and Mitfb) in the development of the zebrafish RPE was explored in these experiments. The loss of Otx activity through morpholino knockdown produced variable eye defects, ranging from delayed RPE pigmentation to severe coloboma, depending on the combination of Otx factors that were targeted. Expression analysis through in situ hybridization demonstrates that <em>otx</em> transcription factors are necessary for the proper expression of <em>mitfa</em> and <em>mitfb</em> while Mitf transcription factors are not required for the expression of <em>otx</em> genes. Surprisingly, the loss of Mitf activity in <em>mitfa</em>, <em>mitfb</em>, or double <em>mitf</em> mutant zebrafish had no effect on RPE pigmentation or development. Moreover, histological analysis revealed that retinal lamination is unaffected in <em>mitf</em> mutants, as well as in <em>otx</em> morphants, even in regions lacking RPE. Otx and Mitf combined loss of function experiments suggest that <em>mitfa</em> and <em>mitfb</em> may still influence zebrafish RPE development. This is further supported by the ability of <em>mitfa</em> to induce pigmentation in the zebrafish retina when misexpressed. These findings suggest that one or more Otx targets in addition to <em>mitfa</em> and <em>mitfb</em>, possibly another <em>mitf</em> family member, are necessary for development of the RPE in zebrafish.</p>