Dataset Information

PRDM16 expression and function in mammalian cochlear development.

ABSTRACT:

Background

PR domain containing 16 (PRDM16) is a key transcriptional regulator in the development of craniofacial, adipose, and neural tissues. Our lab identified PRDM16 expression in the epithelial cells of the Kölliker's organ (KO) that starts at ~E13.5 and is maintained until KO disappearance. A transgenic mouse model that carries a gene trap null allele of Prdm16 (Prdm16^cGT ) was used to characterize the impact of Prdm16 loss on cochlear development.

Results

At P0 Prdm16^cGT null cochlea exhibited hypoplastic KO, shortened cochlear duct, increased density of hair cells (HCs) and supporting cells (SCs) in the apical turn as well as multiple isolated ectopic HCs within the KO domain. KO epithelial cells proliferation rate was reduced in the apical turn of the developing Prdm16^cGT null cochlea vs controls. Bulk RNA sequencing of cochlear duct cells at E14.5 followed by quantitative real time PCR and mRNA Fluorescence in-situ hybridization (FISH) validation identified differentially expressed genes in Prdm16^cGT null vs littermate control cochleae. Upregulated genes at E14.5 included Fgf20, as well as several Notch pathway genes (Lfng, Hes1, and Jag1).

Conclusions

This study characterizes Prdm16 expression during cochlear development and establishes its requirement for KO development.

SUBMITTER: Ebeid M

PROVIDER: S-EPMC9790675 | biostudies-literature | 2022 Oct

REPOSITORIES: biostudies-literature

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Publications

PRDM16 expression and function in mammalian cochlear development.

Ebeid Michael M Barnas Kathy K Zhang Hongji H Yaghmour Amal A Noreikaite Gabriele G Bjork Bryan C BC

Developmental dynamics : an official publication of the American Association of Anatomists 20220507 10

<h4>Background</h4>PR domain containing 16 (PRDM16) is a key transcriptional regulator in the development of craniofacial, adipose, and neural tissues. Our lab identified PRDM16 expression in the epithelial cells of the Kölliker's organ (KO) that starts at ~E13.5 and is maintained until KO disappearance. A transgenic mouse model that carries a gene trap null allele of Prdm16 (Prdm16<sup>cGT</sup> ) was used to characterize the impact of Prdm16 loss on cochlear development.<h4>Results</h4>At P0 P ...[more]

PMID: 35451126

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Project description:BackgroundHearing loss, a major public health issue, affects 1.33 per 1,000 live births worldwide. Genetic factors contribute to over half of congenital cases, with X-linked inheritance accounting for 1-5%. POU3F4 mutations are associated with approximately 50% of X-linked non-syndrome hearing loss cases. POU3F4 plays a critical role in cochlear development by regulating otic mesenchyme cell differentiation. The study investigates the impact of a novel POU3F4 p.E294G mutation on cochlear structure and function using cellular and animal model.MethodsThe study utilized immortalized lymphoblastoid cell lines, POU3F4 overexpressed HEK293 cells and generated Pou3f4 knock-in (Pou3f4KI) mice via CRISPR/Cas9 to introduce the p.E294G mutation. Alterations in expression and subcellular localization of POU3F4 were detected at the cellular level. Auditory function was assessed using auditory brainstem response testing. Cochlear structure was analyzed through histology, immunohistochemistry, scanning electron microscopy, and transmission electron microscopy. RNA sequencing, qPCR and Western blot were conducted to evaluate gene expression and mitochondrial function.ResultsThe transcription of POU3F4 was abnormal and the expression was normal in lymphoblastoid cell lines. Abnormal nuclear localization of POU3F4 p.E294G was found in overexpressed HEK293 cells. Pou3f4KI mice exhibited cochlear malformations, including modiolus hypoplasia and reduced stria vascularis cell populations. Auditory testing revealed progressive hearing loss. Pou3f4 affect mitochondrial protein expression by affecting the expression of TFAM. Mitochondrial dysfunction was evident, with reduced oxidative phosphorylation (OXPHOS) complex assembly and activity, decreased ATP levels. The level of reactive oxygen species, mitochondrial fission and apoptosis in cochlea were elevated.ConclusionsThe POU3F4 p.E294G resulted in abnormal nuclear localization. Pou3f4 mutant disrupts cochlear development and function, impairs mitochondrial integrity, induces oxidative stress, and promotes apoptosis, leading to progressive hearing loss. The findings enhance the understanding of POU3F4-related hearing loss mechanisms and highlight the importance of early genetic screening and audiological monitoring.

Project description:Hair cells of the developing mammalian inner ear are progressively defined through cell fate restriction. This process culminates in the expression of the bHLH transcription factor Atoh1, which is necessary for differentiation of hair cells, but not for their specification. Loss of several genes will disrupt ear morphogenesis or arrest of neurosensory epithelia development. We previously showed in null mutants that the loss of the transcription factor, Gata3, results specifically in the loss of all cochlear neurosensory development. Temporal expression of Gata3 is broad from the otic placode stage through the postnatal ear. It therefore remains unclear at which stage in development Gata3 exerts its effect. To better understand the stage specific effects of Gata3, we investigated the role of Gata3 in cochlear neurosensory specification and differentiation utilizing a LoxP targeted Gata3 line and two Cre lines. Foxg1(Cre)∶Gata3(f/f) mice show recombination of Gata3 around E8.5 but continue to develop a cochlear duct without differentiated hair cells and spiral ganglion neurons. qRT-PCR data show that Atoh1 was down-regulated but not absent in the duct whereas other hair cell specific genes such as Pou4f3 were completely absent. In addition, while Sox2 levels were lower in the Foxg1(Cre):Gata3(f/f) cochlea, Eya1 levels remained normal. We conclude that Eya1 is unable to fully upregulate Atoh1 or Pou4f3, and drive differentiation of hair cells without Gata3. Pax2-Cre∶Gata3(f/f) mice show a delayed recombination of Gata3 in the ear relative to Foxg1(Cre):Gata3(f/f) . These mice exhibited a cochlear duct containing patches of partially differentiated hair cells and developed only few and incorrectly projecting spiral ganglion neurons. Our conditional deletion studies reveal a major role of Gata3 in the signaling of prosensory genes and in the differentiation of cochlear neurosenory cells. We suggest that Gata3 may act in combination with Eya1, Six1, and Sox2 in cochlear prosensory gene signaling.

Dataset Information

PRDM16 expression and function in mammalian cochlear development.

Background

Results

Conclusions

Publications

PRDM16 expression and function in mammalian cochlear development.

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