RT Journal Article
SR Electronic
T1 Identification of novel inhibitors of DLK palmitoylation by High Content Screening
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 430629
DO 10.1101/430629
A1 Dale D.O. Martin
A1 Prasad S. Kanuparthi
A1 Sabrina M. Holland
A1 Shaun S. Sanders
A1 Hey-Kyeong Jeong
A1 Marget B. Einarson
A1 Marlene Jacobson
A1 Gareth M. Thomas
YR 2018
UL http://biorxiv.org/content/early/2018/09/29/430629.abstract
AB After axonal insult and injury, Dual leucine-zipper kinase (DLK) conveys retrograde pro-degenerative signals to neuronal cell bodies via its downstream target c-Jun N-terminal kinase (JNK). We recently reported that such signals critically require modification of DLK by the fatty acid palmitate, via a process called palmitoylation. Compounds that inhibit DLK palmitoylation could thus reduce neurodegeneration, but identifying such inhibitors requires a suitable assay. Here we report that DLK subcellular localization in non-neuronal cells is highly palmitoylation-dependent and can be used as a proxy readout to identify inhibitors of DLK palmitoylation by High Content Screening (HCS). We exploited this highly specific localization of DLK-GFP as the basis for a screen of the Prestwick Compound Library™. We found that ketoconazole, a Prestwick Library compound that most dramatically affected DLK subcellular localization in our primary screen, inhibited DLK palmitoylation in a dose-dependent manner in follow-up biochemical assays. Moeroever, ketoconazole significantly blunted phosphorylation of c-Jun in primary sensory neurons subjected to Trophic Deprivation, a well known model of DLK-dependent pro-degenerative signaling. These findings suggest that our HCS platform is capable of identifying novel inhibitors of DLK palmitoylation and signalling that may have considerable therapeutic potential.