The chaperonin TRiC component Cct3 is required for axonal transport, myelination, and neuromuscular junction...

TRiCopathies are recently discovered neurodevelopmental diseases caused by pathogenic variants in components of the chaperonin tailless complex polypeptide 1 ring complex (TRiC). Composed of chaperonin containing TCP1 subunits 1-8 (CCT1-8), TRiC acts as a chaperone that is required for folding of 10% of proteome, including actin and tubulin. Patients with TRiCopathies display variable combinations of cognitive impairment, epilepsy, polymicrogyria, white matter reduction, cerebellar hypoplasia and alterations in the peripheral neuromuscular system. Here, we aimed at better understanding the pathophysiological role of Cct3 in neurodevelopment, particularly in myelin formation and the neuromuscular system using zebrafish as a model system. We have generated two CRISPR/Cas9 loss-of-function alleles of the orthologous zebrafish cct3 gene. By combining these alleles with transgenic lines and immunostainings we visualized different cell types and subcellular structures in the nervous system by confocal microscopy. Furthermore, we performed electron microscopy to examine zebrafish in comparison to human patient-derived tissue. We demonstrated that cct3 mutant zebrafish fail to form normal myelin sheaths. This was associated with early apoptotic death of neural crest-derived Schwann cells, which were particularly vulnerable to loss of Cct3 function. In line with the observations in patients, developmental refinement of neuromuscular junctions (NMJ) required Cct3. Furthermore, we observed profound cytoskeletal alterations, in particular of tubulin and microtubules as well as severely disturbed microtubule-dependent axonal transport of organelles in peripheral motor axons. Cct3 displays an essential role in myelination, survival of neural crest-derived cells, NMJ refinement, tubulin and microtubule biology as well as axonal transport. Given that axonal transport is essential for transport of axon-glial and signaling factors shaping the NMJ, we speculate that the essential role of Cct3 in axonal transport is a common denominator for the observed phenotypes. These data enhance our understanding of the conserved role of Cct3/TRiC in the developing nervous system and the pathophysiological mechanisms in TRiCopathies.