Target Genes of Neuron-Restrictive Silencer Factor Are Abnormally Up-Regulated in Human Myotilinopathy

Abstract

Myotilinopathy is a subgroup of myofibrillar myopathies caused by mutations in the myotilin gene in which there is aggregation of abnormal cytoskeletal proteins and ubiquitin. We report here on the accumulation of neuron-related proteins such as ubiquitin carboxy-terminal hydrolase L1 (UCHL1) , synaptosomal-associated protein 25 , synaptophysin , and ␣-internexin in aberrant protein aggregates in myotilinopathy. We have determined that the neuronrestrictive silencer factor (NRSF)/RE1 silencing transcription factor (REST), a transcription factor expressed in non-neuronal tissues repressing the expression of several neuronal genes, is reduced in myotilinopathies. Moreover, NRSF transfection reduces UCHL1, synaptosomal-associated protein 25, synaptophysin, and ␣-internexin mRNA levels in DMS53 cells, whereas short interferring NRSF transfection increases UCHL1 and synaptophysin mRNA levels in U87-MG cells. Chromatin immunoprecipitation assays have shown that NRSF interacts with the UCHL1 promoter in U87-MG and HeLa cells. In silico analysis of the UCHL1 gene promoter sequence using the MatInspector software has predicted three potential neuron-restrictive silencer elements ( 1-8 Mutations in several genes have been identified as causing MFMs: desmin, ␣B-crystallin, selenoprotein N, myotilin, ZASP, and filamin C. 5,7,9 -17 The causes of protein aggregation in MFMs are not fully understood, but our previous studies have shown that impaired protein degradation probably plays a crucial role, as suggested by abnormal expression levels and aberrant localization of several subunits of the proteasome 19S and 26S and by the up-regulation of immunoproteasomal subunits in muscle fibers containing abnormal protein deposits.18 Moreover, protein accumulations are enriched in clusterin and ␥-tubulin, whereas p62 and mutant ubiquitin colocalize with protein aggregates, thus suggesting p62 involvement in protein aggregation and mutant ubiquitin in protein degradation in MFMs. 19,20 Preliminary work in our laboratory identified the presence of ubiquitin carboxy-terminal hydrolase L3 (UCHL3) in normal and diseased muscle, but UCHL1 was also present in the setting of abnormal protein deposits in MFMs. This was unexpected, because UCHL1 is abundant in brain and testis, whereas other members of the UCHL family, but not UCHL1, are expressed in other organs. [21][22][23] UCHLs are enzymes involved in the hydrolysis of polyubiquitin chains to increase the availability of free monomeric ubiquitin to the ubiquitin-proteasome

NRSF is an essential mediator for the neuroprotection of trichostatin A in the MPTP mouse model of Parkinson's disease

“…The outcome of interaction between NRSE and NRSF is silencing BDNF expression (Schoenherr and Anderson, 1995, Zuccato et al, 2007, Hara et al, 2009. Moreover, PD-related genes ubiquitin carboxyl-terminal hydroxylase L1 (UCH-L1) and Synapsin have been verified to be NRSF target genes (Barrachina et al, 2007).…”

Myofibrillar myopathies

“…Additional abnormal accumulation of several other proteins was documented in myotilinopathy and desminopathy. These include phospho-tau, β-amyloid, clusterin, a mutant nonfunctional ubiquitin, the multimeric signal protein p62, glycoxidation and lipoxidation markers, neuronal, inducible and endothelial nitric oxide synthases, superoxide dismutase, neuron-related proteins such as ubiquitin carboxy-terminal hydrolase L1, synaptosomal-associated protein 25, synaptophysin, and α-internexin [11,[17][18][19].…”

Myofibrillar Myopathies and the Z-Disk Associated Proteins

“…Analysis of Myotilin levels in patients compared to control individuals failed to identify a reduction in protein level [10,142] with other studies reporting an increase in Myotilin in some patients [142,165]. This observation leads to the hypothesis that mutations in Myotilin affect its dimerisation or interaction with binding partners, resulting in pathology.…”