Let-7 miRNAs control auditory sensory progenitor behavior in the vertebrate inner ear

Abstract

The evolutionary conserved lethal-7 (let-7) family of microRNAs (miRNAs) is a well-known activator of terminal mitosis and differentiation. Surprisingly, we previously found that overexpression of let-7 miRNAs in the murine auditory organ accelerated the terminal mitosis of auditory sensory progenitors (pro-sensory cells) but failed to stimulate their differentiation into mechano-sensory hair cells (HCs). To further address the role of let-7 miRNAs in auditory sensory differentiation, we conducted gain and loss of function experiments in the developing chicken auditory organ, the basilar papilla (BP). Using a sponge approach, we show that the disruption of let-7 miRNA function in the developing BP delays pro-sensory cell exit and delays differentiation of auditory HCs, revealing that endogenous let-7 miRNAs limit pro-sensory cell self-renewal in the developing BP. However, consistent with the role of let-7 miRNAs in the murine auditory organ, let-7b overexpression in the developing BP delayed HC differentiation, suggesting that too low or too high let-7 miRNA levels disrupt HC differentiation. Furthermore, we provide evidence that the repressive role of let-7 miRNAs in HC differentiation may be due to its targeting of the chromatin remodeler CHD7. Mutation in the human CHD7 gene causes CHARGE syndrome, which amongst others is characterized by inner ear and hearing deficits. Using target prediction algorithms, we uncovered a highly predictive and evolutionary conserved let-7 binding site within the Chd7 transcript. Consistent with being a target of let-7 repression, we demonstrate that let-7b overexpression significantly reduced CHD7 protein expression in to the developing BP. Furthermore, utilizing an inducible let-7g transgenic mouse model, we show that let-7 miRNAs negatively regulate CHD7 protein expression in developing murine cochlear, retinal and brain tissue. CHD7 is dosage dependent and the here described regulation by let-7 miRNAs may be critical to fine tune CHD7 protein levels during sensory and neuronal development.\n\nSIGNIFICANCEThe evolutionary highly conserved let-7 miRNAs are essential for proper timing of cell state transitions during embryogenesis. Even though abundantly expressed in the vertebrate auditory organ, surprisingly little is known about their function in auditory sensory differentiation. Here, we demonstrate that endogenous let-7 miRNAs are essential for limiting auditory sensory progenitor (pro-sensory) cell self-renewal. Furthermore, we find that precocious let-7 miRNAs expression interferes with auditory hair cell differentiation and identify chromatin remodeler CHD7 as a potential target gene of let-7 repressive function in HC differentiation.