Proteasomes remain intact, but show early focal alteration in their composition in a mouse model of amyotrophic lateral sclerosis

Abstract

In amyotrophic lateral sclerosis caused by mutations in Cu/Zn‐superoxide dismutase (SOD1), altered solubility and aggregation of the mutant protein implicates failure of pathways for detecting and catabolizing misfolded proteins. Our previous studies demonstrated early reduction of proteasome‐mediated proteolytic activity in lumbar spinal cord of SOD1G93A transgenic mice, tissue particularly vulnerable to disease. The purpose of this study was to identify any underlying abnormalities in proteasomal structure. In lumbar spinal cord of pre‐symptomatic mice [postnatal day 45 (P45) and P75], normal levels of structural 20S α subunits were incorporated into 20S/26S proteasomes; however, proteasomal complexes separated by native gel electrophoresis showed decreased immunoreactivity with antibodies to β3, a structural subunit of the 20S proteasome core, and β5, the subunit with chymotrypsin‐like activity. This occurred prior to increase in β5i immunoproteasomal subunit. mRNA levels were maintained and no association of mutant SOD1 with proteasomes was identified, implicating post‐transcriptional mechanisms. mRNAs also were maintained in laser captured motor neurons at a later stage of disease (P100) in which multiple 20S proteins are reduced relative to the surrounding neuropil. Increase in detergent‐insoluble, ubiquitinated proteins at P75 provided further evidence of stress on mechanisms of protein quality control in multiple cell types prior to significant motor neuron death.

A novel small molecule HSP90 inhibitor, NXD30001, differentially induces heat shock proteins in nervous tissue in culture and in vivo

“…Mutations are thought to promote protein misfolding and inappropriate interactions with multiple macromolecules. Protein chaperoning activity, reflecting the cellular ability to maintain protein conformation, (Bruening et al 1999;Tummala et al 2005) and proteasome activity, reflecting reserve capacity to catabolize misfolded proteins, (Kabashi et al 2004;Cheroni et al 2005Cheroni et al , 2009Kabashi et al 2008) are preferentially compromised in vulnerable regions of spinal cord in SOD1 G93A transgenic mice, a model of fALS due to SOD1 mutation. Similar proteasomal abnormalities were identified in postmortem spinal cord tissue from patients with sALS (Kabashi et al 2012).…”

Functional alterations of the ubiquitin-proteasome system in motor neurons of a mouse model of familial amyotrophic lateral sclerosis†

“…We previously showed a marked decline of constitutive 20S and 19S subunits in motor neurons of symptomatic SOD1G93A mice (10), in line with the decreased catalytic activity of 20S proteasome reported in lumbar spinal cord homogenates of the same mouse model (12). A pre-symptomatic reduction of the 20S constitutive catalytic b3 and b5 subunits without changes in their mRNAs was also reported (11). In the present study, we also found no changes in 20S b5 mRNA levels in the lumbar spinal cord of pre-symptomatic and symptomatic SOD1G93A mice, whereas a moderate decrease in the mRNA levels of the constitutive 19S and 20S a5 occurred at these stages of the disease, suggesting a control at the transcriptional level of these subunits.…”

“…A role of immunoproteasome in ALS has been suggested by the increase in inducible subunit mRNA and protein levels in spinal cord homogenates of rodent models of the pathology (10,11,25,26). At the cellular level, however, the increased expression of immunoproteasome LMP7 (the only subunit investigated) has been principally attributed to glial cells (25,26).…”

“…The replacement of the constitutive subunits with inducible subunits did not result in detectable changes in the 20S proteasome proteolytic activity. Other studies have demonstrated opposite changes in the expression of constitutive and inducible proteasome subunits in SOD1 mutant mice (11,25,26). So far it has not been clarified which mechanisms underlie the shift from constitutive proteasome to immunoproteasome and whether this effect may be related to the pathogenesis and/or disease progression.…”

“…It has been proposed that alterations in the functionality of the ubiquitin-proteasome system (UPS) might play a role in this phenomenon (10)(11)(12)(13)(14)(15).…”