Ubiquitin ligase activity inhibits Cdk5 to control axon termination.

The Cdk5 kinase plays prominent roles in nervous system development, plasticity, behavior and disease. It also has important, non-neuronal functions in cancer, the immune system and insulin secretion. At present, we do not fully understand negative regulatory mechanisms that restrict Cdk5. Here, we use Caenorhabditis elegans to show that CDK-5 is inhibited by the RPM-1/FSN-1 ubiquitin ligase complex. This atypical RING ubiquitin ligase is conserved from C. elegans through mammals. Our finding originated from unbiased, in vivo affinity purification proteomics, which identified CDK-5 as a putative RPM-1 substrate. CRISPR-based, native biochemistry showed that CDK-5 interacts with the RPM-1/FSN-1 ubiquitin ligase complex. A CRISPR engineered RPM-1 substrate 'trap' enriched CDK-5 binding, which was mediated by the FSN-1 substrate recognition module. To test the functional genetic relationship between the RPM-1/FSN-1 ubiquitin ligase complex and CDK-5, we evaluated axon termination in mechanosensory neurons and motor neurons. Our results indicate that RPM-1/FSN-1 ubiquitin ligase activity restricts CDK-5 to control axon termination. Collectively, these proteomic, biochemical and genetic results increase our understanding of mechanisms that restrain Cdk5 in the nervous system.

Axon development is regulated at genetic and proteomic interfaces between the integrin adhesome and the RPM-1 ubiquitin ligase signaling hub.

Integrin signaling plays important roles in development and disease. An adhesion signaling network called the integrin adhesome has been principally defined using bioinformatics and proteomics. To date, the adhesome has not been studied using integrated proteomic and genetic approaches. Here, proteomic studies in C. elegans identified physical associations between the RPM-1 ubiquitin ligase signaling hub and numerous adhesome components including Talin, Kindlin and beta-integrin. C. elegans RPM-1 is orthologous to human MYCBP2, a prominent player in nervous system development associated with a neurodevelopmental disorder. Using neuron-specific, CRISPR loss-of-function strategies, we show that core adhesome components affect axon development and interact genetically with RPM-1. Mechanistically, Talin opposes RPM-1 in a functional 'tug-of-war' on growth cones that is required for accurate axon termination. Thus, our findings orthogonally validate the adhesome via multi-component genetic and physical interfaces with a key neuronal signaling hub and identify new links between the adhesome and brain disorders.