C. elegans DLK-1 mitogen-activated protein kinase kinase kinase (MAPKKK) is the ortholog of the vertebrate MAP3K12 and MAP3K13. This family of MAPKKKs act upstream of MAPKKs to control
p38 or JNK kinase. Recent evidence from multiple animal models have revealed their pivotal roles in stress-sensing, particularly under traumatic injury to the nervous system. In C. elegans, activation of DLK-1 is essential for axon regeneration in multiple types of neurons (Yan et al., 2009, Hammarlund et al., 2009). While it is well established that DLK-1 protein abundance is negatively regulated by the E3 ligase consisted of RPM-1 and FSN-1, it remains possible that there are other mechanisms regulating DLK-1. To address this, we generated a GFP knock-in to tag endogenous DLK-1. We observed that endogenously expressed GFP::DLK-1 is barely detectable, and that in
rpm-1 mutants, the intensity of GFP::DLK-1 in the nervous system is increased, validating the functionality of tagged DLK-1. We have performed visual genetic screens targeting novel regulators of DLK-1 expression. Preliminary characterization of these mutants has revealed multiple phenotypic classes. One class of mutants show ectopic expression of DLK-1 in non-neuronal tissues at early stages, suggesting tissue-specific regulation of
dlk-1. Another class of mutants displayed increased DLK-1 in nearly all neurons, independent of RPM-1. A third class of mutants show up-regulation of DLK-1 in ciliary neurons; and we have determined one causative mutation to be a component of intraflagellar transport complex. Additionally, we identified an intragenic loss of function mutation in DLK-1 kinase domain that causes high level of DLK-1 expression, suggesting that DLK-1 kinase activity is critical for its stability. Taken together, our findings highlight multiple mechanisms controlling DLK-1 with cell-type specificity.