Inhibition of the autophagic protein ULK1 attenuates axonal degeneration in vitro and in vivo and modulates differential splicing

ABSTRACT: Several traumatic and neurodegenerative disorders of the CNS show axonal degeneration as a key and early pathological feature. Following a focal axonal lesion, an extended axonal disintegration by acute axonal degeneration (AAD) occurs within several hours. During AAD, activation of autophagic proteins, including Unc-51 like autophagy activating kinase 1 (ULK1), has been shown but its role is incompletely understood. Here, we overexpressed a dominant negative of ULK1 (ULK1.DN) in primary rat cortical neurons in vitro and in the spinal cord (SC) and optic nerve (ON) in vivo. We found a significant inhibitory effect on autophagy in vitro and in vivo. While no effect on cell survival could be observed, ULK1.DN attenuated AAD after axotomy in microfluidic chambers in vitro. Models of SC injury and ON crush in vivo confirmed an attenuation of axonal degeneration by ULK1.DN. In a translational approach, we employed the selective ULK1 inhibitor SBI-0206965 (SBI) and found an attenuation of AAD after ON crush in vivo mediated by a reduction of autophagy by SBI. Quantitative proteomic profiling after ULK1.DN overexpression in vitro revealed a regulation of proteins associated with translation and splicing. A differential exon expression analysis then identified the genes kinesin family member 1B (Kif1b), DNA damage inducible transcript 3 (Ddit3) and WD repeat domain 6 (Wdr6) as targets of ULK1.DN and potential mediators of its degeneration-attenuating effect. These findings reveal ULK1 as an important mediator of axonal degeneration and elucidate its function in splicing, defining it as a putative therapeutic target.

INSTRUMENT(S): TripleTOF 5600

ORGANISM(S): Rattus Norvegicus (rat)

TISSUE(S): Cell Culture

SUBMITTER: Christof Lenz  

LAB HEAD: Christof Lenz

PROVIDER: PXD011862 | Pride | 2020-04-30

REPOSITORIES: Pride