SkpA Restrains Synaptic Terminal Growth during Development and Promotes Axonal Degeneration following Injury

Abstract

The Wallenda (Wnd)/dual leucine zipper kinase (DLK)-Jnk pathway is an evolutionarily conserved MAPK signaling pathway that functions during neuronal development and following axonal injury. Improper pathway activation causes defects in axonal guidance and synaptic growth, whereas loss-of-function mutations in pathway components impairs axonal regeneration and degeneration after injury. Regulation of this pathway is in part through the E3 ubiquitin ligase Highwire (Hiw), which targets Wnd/DLK for degradation to limit MAPK signaling. To explore mechanisms controlling Wnd/DLK signaling, we performed a large-scale genetic screen in Drosophila to identify negative regulators of the pathway. Here we describe the identification and characterization of SkpA, a core component of SCF E3 ubiquitin ligases. Mutants in SkpA display synaptic overgrowth and an increase in Jnk signaling, similar to hiw mutants. The combination of hypomorphic alleles of SkpA and hiw leads to enhanced synaptic growth. Mutants in the Wnd-Jnk pathway suppress the overgrowth of SkpA mutants demonstrating that the synaptic overgrowth is due to increased Jnk signaling. These findings support the model that SkpA and the E3 ligase Hiw function as part of an SCF-like complex that attenuates Wnd/DLK signaling. In addition, SkpA, like Hiw, is required for synaptic and axonal responses to injury. Synapses in SkpA mutants are more stable following genetic or traumatic axonal injury, and axon loss is delayed in SkpA mutants after nerve crush. As in highwire mutants, this axonal protection requires Nmnat. Hence, SkpA is a novel negative regulator of the Wnd-Jnk pathway that functions with Hiw to regulate both synaptic development and axonal maintenance.

Transmission, Development, and Plasticity of Synapses

“…E3 ubiquitin ligases can act alone or in multicomponent E3 complexes. Recent studies have identified the proteins DFsn, Rae1, and SkpA, which act in a complex with Hiw to regulate Wnd (Wu et al 2007;Tian et al 2011;Brace et al 2014) and resemble the conserved SCF (Skp/Cullin/F-box) E3 complex.…”

PAM forms an atypical SCF ubiquitin ligase complex that ubiquitinates and degrades NMNAT2

“…Proteomics and genetics in flies, as well as biochemistry in 293 cells has suggested PHR proteins can form non-canonical SCF complexes, which contain SKP1 but lack a Cullin (37,40). While these studies provided evidence individual components can interact, biochemistry showing that FBXO45 and SKP1, but not CUL1, assemble into a complex with PAM remains notably absent.…”

Defining Minimal Binding Regions in Regulator of Presynaptic Morphology 1 (RPM-1) Using Caenorhabditis elegans Neurons Reveals Differential Signaling Complexes

“…In C. elegans, RPM-1 functions in an Skp/Cullin/F-box (SCF) complex that includes the F-box protein FSN-1, the Skp protein SKR-1, and the Cullin CUL-1 (20). In mammals and flies, Phr1 and Highwire function in a non-canonical SCF complex that contains the F-box protein Fbxo45 and Skp1 but lacks a Cullin (21)(22)(23). Despite differences in the composition of PHR ubiquitin ligase complexes, the functional relationship between PHR proteins and FSN1⅐Fbxo45 is evolutionarily conserved.…”