Defining tissue-specific developmental requirements for the proteasome

Abstract

The ubiquitin-proteasome system provides a rapid, temporally controlled, and highly specific mechanism for intracellular protein degradation. As part of its role in degrading short-lived regulatory proteins, the proteasome regulates multiple cellular processes integral to embryonic development. However, time- and tissue-specific requirements for the proteasome during development are currently not well understood, as proteasome subunit mutations are frequently early embryonic lethal. Here I leverage zebrafish mutants in the 20S proteasome subunit psmb1 and the 19S subunit psmc6 to examine developmental phenotypes arising from loss of proteasome function. Deep phenotyping of psmb1 and psmc6 mutants revealed striking defects in craniofacial cartilage development, impaired hepatopancreatic development, abnormalities in the cranial ganglia, and increased apoptosis in the brain and spinal cord. In contrast, psmb1 was largely dispensable for kidney and heart development, underscoring varying tissue-sensitivity to loss of proteasome function. To elucidate the cellular mechanisms linking proteasome inhibition and craniofacial dysmorphisms, I performed time-lapse imaging of craniofacial cartilage, muscle, and tendon development in combination with transcriptomic analysis of craniofacial chondrocytes and muscle in psmb1 mutants. I found that psmb1 is required for cartilage, muscle, and tendon morphogenesis, and that maturation of craniofacial cartilage and muscle is also compromised. Overexpression of psmb1 in neural crest cells and chondrocytes rescued the cartilage and tendon phenotypes, but induced only a partial rescue of the muscle, demonstrating that psmb1 is required in both tissue-autonomous and non-autonomous fashions during craniofacial development. In sum, this work establishes zebrafish psmb1 and psmc6 mutants as essential models to investigate the mechanisms linking proteasome mutations with specific developmental phenotypes and identifies the cellular mechanisms through which proteasome mutations induce craniofacial malformations.