Temporal and spatial assembly of inner ear hair cell ankle link condensate through phase separation

Abstract

Stereocilia are actin-based cell protrusions of inner ear hair cells and are indispensable for mechanotransduction. Ankle links connect the ankle region of developing stereocilia, playing an essential role in stereocilia development. WHRN, PDZD7, ADGRV1 and USH2A have been identified to form the so-called ankle link complex (ALC); however, the detailed mechanism underlying the temporal emergence and degeneration of ankle links remains elusive. Here we show that WHRN and PDZD7 orchestrate ADGRV1 and USH2A to assemble the ALC through liquid-liquid phase separation (LLPS). Disruption of the ALC multivalency for LLPS largely abolishes the distribution of WHRN at the ankle region of stereocilia. Interestingly, high concentration of ADGRV1 inhibits LLPS, providing a potential mechanism for ALC disassembly. Moreover, certain deafness mutations of ALC genes weaken the multivalent interactions of ALC and impair LLPS. In conclusion, our study demonstrates that LLPS mediates ALC formation, providing essential clues for understanding the pathogenesis of deafness.

Usher syndrome proteins ADGRV1 (USH2C) and CIB2 (USH1J) interact and share a common interactome containing TRiC/CCT-BBS chaperonins

“…In addition, we showed the localization of ADGRV1 in mitochondria-associated ER membranes (MAMs), important for the maintenance of Ca 2+ -homeostasis (Krzysko et al, 2022(Krzysko et al, ), 2022 and that ADGRV1 controls autophagy processes and cellular proteostasis (Linnert et al, 2023). Moreover, others and we have previously shown that the USH2C protein ADGRV1 is part of the USH interactome interacting with both other USH2 proteins, usherin (USH2A) and whirlin (USH2D), and the two USH1 proteins myosin VIIa (USH1B) and harmonin (USH1C) (Reiners et al, 2005;van Wijk et al, 2006;Michalski et al, 2007;Wang et al, 2023). The putative USH1J protein CIB2 also interacts with the USH proteins whirlin and myosin VIIa (Riazuddin et al, 2012), and the interaction between ADGRV1 and CIB2 that we describe here confirms that CIB2 is part of the USH interactome.…”

Decoding Missense Variants by Incorporating Phase Separation via Machine Learning

“…Aberrant phase separation (PS) or aggregation processes have been implicated in the pathogenesis of various diseases, including neurodegenerative disorders, autism 4 , and hearing loss [7][8][9] . In this study, we employed PSMutPred to predict the PS impact ('Impact' prediction and 'Strengthen/Weaken' prediction) of missense variants from genes associated with PS-related diseases.…”

“…An increasing number of proteins are recognized for their roles via the formation of phase separation condensates. These condensates include the nucleolus and transcription factories in the nucleus 4 , stress granules 5 , and protein densities at neuronal synapses 6 and inner-ear hair cells [7][8][9] .…”