Mutant Cu/Zn-Superoxide Dismutase Associated with Amyotrophic Lateral Sclerosis Destabilizes Vascular Endothelial Growth Factor mRNA and Downregulates Its Expression

Abstract

Vascular endothelial growth factor (VEGF) plays a neuroprotective role in mice harboring mutations of copper-zinc superoxide dismutase 1 (SOD1) in familial amyotrophic lateral sclerosis (ALS). Conversely, the loss of VEGF expression through genetic depletion can give rise to a phenotype resembling ALS independent of SOD1 mutations. Here, we observe a profound downregulation of VEGF mRNA expression in spinal cords of G93A SOD1 mice that occurred early in the course of the disease. Using an in vitro culture model of glial cells expressing mutant SOD1, we demonstrate destabilization and downregulation of VEGF RNA with concomitant loss of protein expression that correlates with level of transgene expression. Using a luciferase reporter assay, we show that this molecular effect is mediated through a portion of the VEGF 3Ј-untranslated region (UTR) that harbors a class II adenylate/uridylate-rich element. Other mutant forms of SOD1 produced a similar negative effect on luciferase RNA and protein expression. Mobility shift assay with a VEGF 3Ј-UTR probe reveals an aberrantly migrating complex that contains mutant SOD1. We further show that the RNA stabilizing protein, HuR (human antigen R), is translocated from nucleus to cytoplasm in mutant SOD1 cells in vitro and mouse motor neurons in vivo. In summary, our data suggest that mutant SOD1 gains a novel function, possibly by altering the ribonucleoprotein complex with the VEGF 3Ј-UTR. We postulate that the resultant dysregulation of VEGF posttranscriptional processing critically reduces the level of this neuroprotective growth factor and accelerates the neurodegenerative process in ALS.

Intrathecal transplantation of human neural stem cells overexpressing VEGF provide behavioral improvement, disease onset delay and survival extension in transgenic ALS mice

“…38,[41][42][43] Of these neurotrophic factors, VEGF is well known to have an important function in the pathogenesis of ALS. [31][32][33][34]37,38 The genetically modified human NSCs we established, F3.VEGF cells, were confirmed to release four times of VEGF165, the most abundant and biologically active isoform in vivo, compared with the parental F3 cells as we have reported previously in animal models of stroke. 28 Ex vivo gene therapy by genetically modified cells to carry growth factor gene has no risk related with virus injection during the in vivo gene therapy, and the transplanted cells can deliver growth factor over a large area as the cells migrate and are integrated to the host brain tissue.…”

“…37 In SOD mutant mice, a significant downregulation of VEGF mRNA expression in spinal cord was found early in the course of the disease indicating that dysregulation of VEGF posttranscriptional processing reduces VEGF levels. 31 Subsequent follow-up human genetic studies have shown that 'low-VEGF' haplotypes in the VEGF promoter had an increased risk of ALS in the European population. 38 Several reports have also reported low VEGF levels in the colony-stimulating factor of ALS patients, [32][33][34] and the expression of both VEGF and VEGFR2 was downregulated on the anterior horn motoneurons as compared with the normal controls.…”

“…The expression of mutant SOD1 protein in ALS animals destabilizes VEGF mRNA and markedly downregulates protein production, and this dysregulation of VEGF posttranscriptional processing critically reduces the level of neuroprotective VEGF and accelerates the neurodegenerative process in ALS. 31 In fact, the VEGF levels in cerebrospinal fluid tended to be lower in ALS patients than in the normal population. [32][33][34] Considering the evidence of clinical improvement in ALS animal models after transplantation of stem cells [10][11][12][13] and VEGF treatment, [16][17][18][19]29,30 we wished to investigate whether the human NSCs overexpressing VEGF, by pairing clonal human NSCs with VEGF gene, can lead to the clinical improvement in SOD1G93A mouse model of ALS.…”

SOD1 Function and Its Implications for Amyotrophic Lateral Sclerosis Pathology

“…These interactions, which are specific to mutant SOD1, negatively affect levels of VEGF mRNA, which is a neuroprotective factor for motor neurons (Lu and others 2007;Lu and others 2009). Similarly, mutant SOD1 has been shown to bind the 3′ UTR of the neurofilament light chain (NFL) mRNA and negatively affect its stability.…”

“…Whilst evidence of a role for SOD1 in the former two is still lacking, mutant SOD1 can bind mRNAs and play a role in their stabilization (Lu and others 2007;Lu and others 2009;Chen and others 2014). Wildtype SOD1 has further been shown to interact with TDP43, suggesting a potential common action in the regulation of specific RNA stability (Volkening and others 2009).…”