Imaging of Sensory Hair Cells in the Cochlear Hearing Organ by Cryo Soft X-ray Tomography

Our project focuses on a protein family implicated in Usher syndrome, the most common inherited disorder causing combined deafness and blindness. Mutations in genes encoding Usher proteins severely impair sensory cells responsible for hearing, vision, and balance. We aim to uncover the molecular and cellular mechanisms by which the Usher 2 complex contributes to stereocilia bundle development at the apex of cochlear hair cells.

In the cochlea, the organ of hearing, hair cells are arranged in four rows, each bearing highly organized stereocilia. The Usher 2 complex, essential for cochlear development, involves scaffolding proteins at the base of these bundles. Our objective is to determine the in situ 3D architecture of hair bundles and the native Usher 2 complex, and to assess how deafness-causing mutations disrupt its organization.

We have proposed to combine cutting edge imaging methods: cryo-electron tomography (cryoET), cryo-soft X-ray tomography (cryoSXT), and super-resolution fluorescence nanoscopy (STED). These approaches will provide structural data from the microscale (hair cells) down to the nanoscale and atomic scale (protein domains). Combining in situ reconstructions with high-resolution structures of Usher proteins from NMR and single-particle cryoEM will enable us to build a detailed model of Usher 2 protein organization and interactions in their native environment.This structural characterization will advance our understanding of the physiopathology of Usher syndrome type 2 and could inform improved diagnostic approaches.

The project is part of the Horizon Europe MSCA doctoral network CLEXM and will be conducted by PhD student Antonio El Helou, co-supervised by Nicolas Wolff and Anna Sartori-Rupp (Institut Pasteur, Paris, France). Using nanometric-resolution reconstructions of mouse cochlear stereocilia, we aim to elucidate how Usher proteins form a protein network that drives hair bundle growth and morphogenesis essential for hearing.