Biology subjects

Tavares, A. L. P.

Publications and source records attributed to Tavares, A. L. P..

3 recordsLinked to original sources

bioRxiv2021

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 Meckels, 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.

developmental biology↗

bioRxiv2021

Mutations in SIX1 associated with Branchio-oto-renal Syndrome (BOR) differentially affect otic expression of putative target genes

Single nucleotide mutations in SIX1 are causative in some individuals diagnosed with branchio-otic/branchio-oto-renal (BOR) syndrome. To test whether these mutations have differential effects on otic gene expression, we engineered four BOR mutations in Xenopus six1 and targeted mutant protein expression to the neural crest and cranial placode precursor cells in wild-type embryos. Changes in the otic expression of putative Six1 targets and/or co-factors were monitored by qRT-PCR and in situ hybridization. We found that each mutant had a different combination of effects. The V17E mutant reduced eya2, tspan13, zbtb16 and pa2g4 otic vesicle expression at a frequency indistinguishable from wild-type Six1, but reduced prdm1 more and spry1 less compared to wild-type Six1. For most of these genes, the R110W, W122R and Y129C mutants were significantly less repressive compared to wild-type Six1. Their individual effects varied according to the level at which they were expressed. The R110W, W122R and Y129C mutants also often expanded prdm1 otic expression. Since previous studies showed that all four mutants are transcriptionally deficient and differ in their ability to interact with co-factors such as Eya1, we propose that altered co-factor interactions at the mutated sites differentially interfere with their ability to drive otic gene expression.

developmental biology↗

bioRxiv2021

Sobp modulates Six1 transcriptional activation and is required during craniofacial development

Branchio-oto-renal syndrome (BOR) is a disorder characterized by hearing loss, craniofacial and/or renal defects. Mutations in the transcription factor Six1 and its cofactor Eya1, both required for otic development, are linked to BOR. We previously identified Sobp as a potential Six1 cofactor and SOBP mutations in mouse and humans cause otic phenotypes; therefore, we asked whether Sobp interacts with Six1 and thereby may contribute to BOR. Co-IP and immunofluorescence experiments demonstrate that Sobp binds to and co-localizes with Six1 in the cell nucleus. Luciferase assays show that Sobp represses Six1+Eya1 transcriptional activation. Experiments in Xenopus embryos that either knockdown or increase expression show that Sobp is required for formation of ectodermal domains at neural plate stages. In addition, altering Sobp levels disrupts otic vesicle development and causes craniofacial cartilage defects. Expression of Xenopus Sobp containing the human mutation disrupts the pre-placodal ectoderm similar to full-length Sobp, but other changes are distinct. These results indicate that Sobp modifies Six1 function, is required for vertebrate craniofacial development, and identifies Sobp as a potential candidate gene for BOR and other deafness syndromes. Summary statementSobp interacts with Six1 in the cell nucleus and represses the Six1+Eya1 transcriptional activation. In Xenopus embryos, Sobp functions during early stages of inner ear development.

developmental biology↗