ldn-57444 and Disease-Models--Animal

Levels of ubiquitin carboxyl-terminal hydrolase L1 (UCHL1) are robustly increased in spinal muscular atrophy (SMA) patient fibroblasts and mouse models. We therefore wanted to establish whether changes in UCHL1 contribute directly to disease pathogenesis, and to assess whether pharmacological inhibition of UCHL1 represents a viable therapeutic option for SMA.. SMA mice and control littermates received a pharmacological UCHL1 inhibitor (LDN-57444) or DMSO vehicle. Survival and weight were monitored daily, a righting test of motor performance was performed, and motor neurone loss, muscle fibre atrophy and neuromuscular junction pathology were all quantified. Ubiquitin-like modifier activating enzyme 1 (Uba1) was then pharmacologically inhibited in neurones in vitro to examine the relationship between Uba1 levels and UCHL1 in SMA.. Pharmacological inhibition of UCHL1 failed to improve survival, motor symptoms or neuromuscular pathology in SMA mice and actually precipitated the onset of weight loss. LDN-57444 treatment significantly decreased spinal cord mono-ubiquitin levels, further exacerbating ubiquitination defects in SMA mice. Pharmacological inhibition of Uba1, levels of which are robustly reduced in SMA, was sufficient to induce accumulation of UCHL1 in primary neuronal cultures.. Pharmacological inhibition of UCHL1 exacerbates rather than ameliorates disease symptoms in a mouse model of SMA. Thus, pharmacological inhibition of UCHL1 is not a viable therapeutic target for SMA. Moreover, increased levels of UCHL1 in SMA likely represent a downstream consequence of decreased Uba1 levels, indicative of an attempted supportive compensatory response to defects in ubiquitin homeostasis caused by low levels of SMN protein.

Topics: Animals; Cells, Cultured; Disease Models, Animal; Fibroblasts; Homeostasis; Humans; Indoles; Mice; Motor Activity; Motor Neurons; Muscular Atrophy, Spinal; Oximes; Survival of Motor Neuron 1 Protein; Ubiquitin Thiolesterase

Deposition of amyloid β protein (Aβ) in the brain is the hallmark of Alzheimer's disease (AD) pathogenesis. Beta-site amyloid precursor protein (APP) cleaving enzyme 1 (BACE1) is the β-secretase in vivo essential for generation of Aβ. Previously we demonstrated that BACE1 is ubiquitinated and the degradation of BACE1 is mediated by the ubiquitin-proteasome pathway (UPP). However the mechanism underlying regulation of BACE1 degradation by UPP remains elusive. Ubiquitin carboxyl-terminal hydrolase L1 (UCHL1) is a deubiquitinating enzyme highly specific to neuron, catalyzing the hydrolysis of ubiquitin conjugates from ubiquitinated substrates. UCHL1 regulates ubiquitin-dependent protein degradation. However, whether UCHL1 is particularly involved in the proteasomal degradation of BACE1 and what is the role of UCHL1 in AD pathogenesis remain elusive. To investigate the effect of UCHL1 on BACE1 degradation, HUCH cells, a UCHL1 stably over-expressed HEK293 cell line, was established. We found that inhibition of UCHL1 significantly increased BACE1 protein level in a time-dependent manner. Half life of BACE1 was reduced in HUCH cells compared with HEK. Over-expression of UCHL1 decreased APP C-terminal fragment C99 and Aβ levels in HUCH cells. Moreover, disruption of Uchl1 gene significantly elevated levels of endogenous BACE1, C99 and Aβ in the Uchl1-null gad mice. These results demonstrated that UCHL1 accelerates BACE1 degradation and affects APP processing and Aβ production. This study suggests that potentiation of UCHL1 might be able to reduce the level of BACE1 and Aβ in brain, which makes it a novel target for AD drug development.

Topics: Amyloid beta-Peptides; Amyloid beta-Protein Precursor; Amyloid Precursor Protein Secretases; Animals; Antibodies; Aspartic Acid Endopeptidases; Cell Line, Transformed; Cycloheximide; Disease Models, Animal; Enzyme Inhibitors; Enzyme-Linked Immunosorbent Assay; Gene Expression Regulation; Indoles; Mice; Mice, Neurologic Mutants; Mutation; Neuroaxonal Dystrophies; Oximes; Peptide Fragments; Protein Processing, Post-Translational; Protein Synthesis Inhibitors; Time Factors; Transfection; Ubiquitin Thiolesterase