Interactions of Nesprin-4-Containing LINC Complexes in Outer Hair Cells Explored by BioID

As components of diverse tissues and organs, metazoan cells have to display a wide variety of specialized functions. Implementation of such functions invariably entails the establishment of tissue-specific cellular architecture (Bone and Starr, J Cell Sci 129:1951-1961, 2016). In animal cells, the nucleus is typically the largest organelle and in many respects acts as a landmark for multiple subcellular structures. For instance, in epithelial cells, the nucleus is frequently positioned close to the basal membrane via association with the cytoskeleton. Clearly such associations must be mediated by protein components of the outer nuclear membrane. One such protein is Nesprin-4, a member of the KASH domain family that is expressed in a variety of epithelial cells, including sensory outer hair cells of the inner ear. In this chapter, I describe a proximity-based biotinylation technique, BioID, that can be applied to Nesprin-4 to map its interactions at the nuclear periphery.