The role of tissue transglutaminase in 1-methyl-4-phenylpyridinium (MPP+)-induced toxicity in differentiated human SH-SY5Y neuroblastoma cells

Abstract

Tissue transglutaminase (TG2) can induce post-translational modification of proteins, resulting in protein cross-linking or incorporation of polyamines into substrates, and can also function as a signal transducing G protein. The role of TG2 in the formation of insoluble cross-links has led to its implication in some neurodegenerative conditions. Exposure of pre-differentiated SH-SY5Y cells to the Parkinsonian neurotoxin 1-methyl-4-phenylpyridinium ion (MPP + ) resulted in significant dose-dependent reductions in TG2 protein levels, measured by probing Western blots with a TG2-specific antibody. Transglutaminase (TG) transamidating activity, on the other hand, monitored by incorporation of a polyamine pseudo-substrate into cellular proteins, was increased. Inhibitors of TG (putrescine) and TG2 (R283) exacerbated MPP + toxicity, suggesting that activation of TG2 may promote a survival response in this toxicity paradigm.

Blockade of enzyme activity inhibits tissue transglutaminase-mediated transamidation of α-synuclein in a cellular model of Parkinson's disease

“…Therefore, we treated the cells with the mitochondrial complex I inhibitor MPP + , an active metabolite of 1-methyl-4-phenyl-1,2,3,6 tetrahydropyridine (MPTP) that is commonly used as a toxin to induce PD-like phenomena in several well-established cellular and in vivo PD models (Burns et al, 1983;Heikkila et al, 1984). In line with previous work by Beck et al (2006), we found that incubation with MPP + (1-5 mM) for 24 h induced a concentration-dependent activation of tTG activity in RA-differentiated WT SH-SY5Y cells (Fig. 5A).…”

“…2A) In contrast to the in vitro data, the results from our cellular model clearly distinguished between the different drugs on both potency and efficacy of tTG inhibition. In fact, to our knowledge, our data are among the first to establish a clear concentrationeffect relationship of pharmacologic tTG inhibition in cells, although using a different experimental setup, effects of inhibitors on tTG activity in SH-SY5Y cells have been reported previously (Beck et al, 2006;Lesort et al, 2003;Singh et al, 2003). This model, originally characterized by Zhang et al (1998), is based on detection of tTG-mediated covalent incorporation of the amine donor BAP into cellular proteins following activation of transamidation activity by a rise in intracellular Ca 2+ concentration as a result of exposure of the cells to Ca 2+ ionophores like A23187.…”

“…However, under pathological conditions such as prevail in PD, cytoplasmic Ca 2+ concentration rises to levels which enable tTG to interact with its substrates (Zhang et al, 1998). To specifically identify tTG modified forms of a-synuclein under experimental conditions mimicking PD, we activated intracellular tTG either through stimulation of Ca 2+ influx with the ionophore A23187 (Fesus et al, 1981) or inhibition of mitochondrial respiration with the PD mimetic MPP + (Beck et al, 2006). Simultaneously, we incubated the cells with the well-characterized labelled synthetic tTG substrate polyamine BAP (Zhang et al, 1998), followed by enrichment of BAP-positive asynuclein protein.…”

Lipopolysaccharide-Stimulated Transglutaminase 2 Expression Enhances Endocytosis Activity in the Mouse Microglial Cell Line BV-2

“…Total TG activity was determined using a method developed by Beck et al [12] with modifications. BV-2 cells (8 × 10 5 cells/dish) were plated onto non-coating 35-mm dishes and stimulated with 300 ng/ml LPS for 24 h in the presence of 250 µ M biotinylated pentylamine (Thermo Fisher Scientific, Waltham, Mass., USA) as the substrate for TG enzyme.…”

Microglial endocytosis and transglutaminases

“…It has been reported the association between TGs and neurodegeneration [41][42][43]. On the other hand, microglial phagocytosis has been reported to be involved in neuronal death in neurodegenerative diseases [44,45].…”