The evolutionarily conserved PHR (Pam/Highwire/RPM-1) proteins play distinct roles in axon guidance, axon termination, and synapse formation. Studies in C. elegans, Drosophila, and mouse show that an important function of PHR proteins is to act as E3 ubiquitin ligases for the Dual-Leucine-zipper-bearing MAPKKK (DLK), which results in negative regulation of the DLK MAP kinase cascade. However, the signaling output of the DLK kinase cascade is largely unknown. Previous screens for suppressors of
rpm-1 neuronal defects uncovered many alleles of the MAPK genes
dlk-1,
mkk-4, and
pmk-3 (Nakata et al., 2005). By analyzing additional
rpm-1(lf) suppressors, we identified two new loci. One is a member of the MAP kinase-activated protein kinase family (MAPKAPK), MAK-2, which is closely related to murine MAPKAPK2 (MK2). The other is CEBP-1, a member of the C/EBP subfamily of bZip domain proteins. By biochemical studies, we find MAK-2 is directly phosphorylated by PMK-3. Phosphorylation of the conserved threonine residues is required for MAK-2 rescuing activity. Expression of phosphomimetic mutant forms of MAK-2 induces gain of function phenotypes that are dependent on CEBP-1 but not PMK-3. These data establish that MAK-2 acts downstream of PMK-3 and upstream of CEBP-1. Unexpectedly, functional CEBP-1::GFP protein is present in axons and at synapses,In addition to the nucleus.Using an RNA tagging strategy we find that
cebp-1 mRNAs are also localized to axons and synapses, and that this localization requires the
cebp-1 3'' UTR. We show that activation of the DLK-1 pathway causes stabilization of
cebp-1 mRNAs. The 3'' UTR of
cebp-1 is necessary and sufficient for this stabilization. Inappropriate elevation of
cebp-1 mRNA in adult neurons disrupts synapses and axon morphology, showing that the DLK-1 pathway is required in mature neuron maintenance and synapse plasticity. Furthermore, we find that CEBP-1 and the DLK-1 pathway are essential for axon regeneration after laser axotomy in adult touch neurons. We show that axotomy triggers local synthesis of new CEBP-1 in injured axons. This local synthesis is dependent on DLK-1 and requires the
cebp-1 3''UTR. Our findings identify the DLK-1 pathway as a regulator of mRNA stability in synapse formation and maintenance, and also in adult axon regeneration.