Dual Leucine Zipper Kinase (DLK) Constitutively Signals in the Uninjured Mouse Cerebellum

Abstract

DLK is a master regulator of axonal retraction and regeneration following neuronal injury and contributes to neurodegeneration in numerous animal models of disease, but the mechanisms by which DLK's pro‐degenerative actions are constrained in healthy neurons are not known. Data to this point demonstrates that DLK is constitutively expressed in post‐mitotic neurons where it is maintained in an inactive state. Once injury occurs, DLK protein is stabilized, phosphorylated, and liberated from inhibitory factors, allowing it to engage the mitogen‐activated protein kinase kinase 4 (MKK4) to phospho‐c‐Jun N‐terminal Kinase (JNK) signaling cascade to regulate apoptosis via phospho‐c‐jun (p‐c‐jun) and the integrated stress response (ISR) pathway via protein kinase R (PKR)‐like endoplasmic reticulum kinase (PERK). Here we report that DLK also functions in healthy neurons as it is constitutively active in the adult mouse cerebellum in the absence of injury. Mouse cerebellar granule cells express full‐length DLK protein predominantly in the axon terminals and post‐synaptic densities along with abundant nuclear p‐c‐jun. Pharmacological inhibition of DLK rapidly eliminates nearly all phospho‐Mkk4 and nuclear p‐c‐Jun within 2 hours, demonstrating that c‐jun phosphorylation is DLK dependent. To understand how DLK pathway activation differs between homeostatic signaling and disease response, we used western blotting and immunofluorescence to compare the DLK pathway between cerebellum, cortex, and hippocampus in rTg4510 mice, which express human P301L mutant tau in the forebrain where it leads to profound DLK activation and neurodegeneration, with non‐transgenic controls. p‐c‐jun is significantly higher in cerebellum and in the transgenic forebrain areas than in the non‐transgenic control forebrain confirming that it is constitutively expressed in the cerebellum and induced from lower basal levels by pathological tau. We next examined the PERK pathway, including phospho‐PERK, phospho‐eIF2a, and the induction of GADD34, ATF4, and CHOP. Like p‐c‐jun, PERK signaling was induced in the rTg4510 forebrain, but was not activated to the same extent in the cerebellum, suggesting that DLK signaling during disease involves enhanced PERK signaling. DLK to JNK signaling is orchestrated by the JNK interacting protein (JIP) scaffolding molecules, of which there are 3 members. In contrast to the forebrain, the cerebellum expresses very little JIP3, whereas JIP1 and JIP2 appear similar across regions. Therefore we hypothesize that JIP3 is required for the injury‐ and disease‐induced DLK signaling pathway. Transcriptomics are currently being utilized to elucidate the physiological relevance of constitutive DLK signaling in the cerebellum.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.