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Huntington disease iPSCs show early molecular changes in intracellular signaling, the expression of oxidative stress proteins and the p53 pathway.

Szlachcic WJ, Switonski PM, Krzyzosiak WJ, Figlerowicz M, Figiel M - Dis Model Mech (2015)

Bottom Line: Several molecular and developmental effects of HD have been identified using neural stem cells (NSCs) and differentiated cells, such as neurons and astrocytes.Surprisingly, we found that a number of changes affecting cellular processes in HD were also present in undifferentiated pluripotent HD iPSCs, including the dysregulation of the MAPK and Wnt signaling pathways and the dysregulation of the expression of genes related to oxidative stress, such as Sod1.Interestingly, a common protein interactor of the huntingtin protein and the proteins in the above pathways is p53, and the expression of p53 was dysregulated in HD YAC128 iPSCs and human HD iPSCs.

View Article: PubMed Central - PubMed

Affiliation: Institute of Bioorganic Chemistry, Polish Academy of Sciences, Noskowskiego 12/14, Poznań 61-704, Poland.

No MeSH data available.


Related in: MedlinePlus

The MAPK pathway is dysregulated in the YAC128 iPSCs. (A) Representative time-course of ERK1/2 phosphorylation stimulated by bFGF in the YAC128 and wild-type (WT) iPSC lines. The ES lane represents a standard protein lysate that was used to normalize the band intensity among experiments. (B) Diagram demonstrating the profile of ERK1/2 activation (levels of pERK1/2). The data for the diagram were collected by investigating the level of pERK1/2 in several different clonal lines of HD YAC128 (n=5) and WT iPSCs (n=6). ERK1/2 activation was dramatically decreased in the HD YAC128 iPSCs 10 min after stimulation using bFGF, whereas ERK1/2 activation in the WT iPSCs remained high. Two-way ANOVA; time×genotype interaction, P=0.0013; Bonferroni post-test. (C) Western blot analysis of the levels of total ERK1/2 proteins. (D) The analysis revealed a decreased level of total ERK1 in the YAC128 lines (t-test, P=0.0189). * indicates the YAC128/Oct-eGFP lines.
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DMM019406F3: The MAPK pathway is dysregulated in the YAC128 iPSCs. (A) Representative time-course of ERK1/2 phosphorylation stimulated by bFGF in the YAC128 and wild-type (WT) iPSC lines. The ES lane represents a standard protein lysate that was used to normalize the band intensity among experiments. (B) Diagram demonstrating the profile of ERK1/2 activation (levels of pERK1/2). The data for the diagram were collected by investigating the level of pERK1/2 in several different clonal lines of HD YAC128 (n=5) and WT iPSCs (n=6). ERK1/2 activation was dramatically decreased in the HD YAC128 iPSCs 10 min after stimulation using bFGF, whereas ERK1/2 activation in the WT iPSCs remained high. Two-way ANOVA; time×genotype interaction, P=0.0013; Bonferroni post-test. (C) Western blot analysis of the levels of total ERK1/2 proteins. (D) The analysis revealed a decreased level of total ERK1 in the YAC128 lines (t-test, P=0.0189). * indicates the YAC128/Oct-eGFP lines.

Mentions: The ERK pathway has been implicated in HD pathogenesis (Bowles and Jones, 2014), and upregulation of its activity might be beneficial in mouse HD models (Maher et al., 2011). Therefore, we investigated the profile of ERK activation in YAC128 iPSCs expressing human mutant huntingtin and investigated whether HD cells in an ESC-like state exhibited dysregulation of the ERK pathway. We stimulated both YAC128 and WT iPSCs using bFGF for 5, 10 or 15 min and compared their levels of ERK pathway activation (Fig. 3A). We observed different profiles of induction of this pathway for WT and HD YAC128 iPSCs (time×genotype interaction, P=0.0013; main effect of time, P<0.0001; two-way ANOVA). In WT iPSCs, we observed a maximum of approximately a 29-fold increase in the ERK1/2 phosphorylation at 5 min compared with basal levels, whereas the level of activation observed at 15 min after stimulation had decreased to 11-fold above basal levels. In YAC128 iPSCs, we observed a similar peak of ERK1/2 activation at 5 min after stimulation. However, the activation of ERK1/2 recorded at 10 min after stimulation had decreased 2.3-fold, whereas the level of ERK1/2 activation in WT iPSCs remained unchanged (P<0.01, Bonferroni post-test; Fig. 3B). In addition, we found that the level of basal activation of ERK1/2 in YAC128 iPSCs was 1.56-fold higher than that in WT iPSCs, although this difference was not significant (P=0.08). Moreover, the analyses of the levels of the total ERK1 and 2 proteins revealed that the total ERK1 protein levels were significantly lower in the YAC128-iPSC cells (Fig. 3C,D).Fig. 3.


Huntington disease iPSCs show early molecular changes in intracellular signaling, the expression of oxidative stress proteins and the p53 pathway.

Szlachcic WJ, Switonski PM, Krzyzosiak WJ, Figlerowicz M, Figiel M - Dis Model Mech (2015)

The MAPK pathway is dysregulated in the YAC128 iPSCs. (A) Representative time-course of ERK1/2 phosphorylation stimulated by bFGF in the YAC128 and wild-type (WT) iPSC lines. The ES lane represents a standard protein lysate that was used to normalize the band intensity among experiments. (B) Diagram demonstrating the profile of ERK1/2 activation (levels of pERK1/2). The data for the diagram were collected by investigating the level of pERK1/2 in several different clonal lines of HD YAC128 (n=5) and WT iPSCs (n=6). ERK1/2 activation was dramatically decreased in the HD YAC128 iPSCs 10 min after stimulation using bFGF, whereas ERK1/2 activation in the WT iPSCs remained high. Two-way ANOVA; time×genotype interaction, P=0.0013; Bonferroni post-test. (C) Western blot analysis of the levels of total ERK1/2 proteins. (D) The analysis revealed a decreased level of total ERK1 in the YAC128 lines (t-test, P=0.0189). * indicates the YAC128/Oct-eGFP lines.
© Copyright Policy - open-access
Related In: Results  -  Collection

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DMM019406F3: The MAPK pathway is dysregulated in the YAC128 iPSCs. (A) Representative time-course of ERK1/2 phosphorylation stimulated by bFGF in the YAC128 and wild-type (WT) iPSC lines. The ES lane represents a standard protein lysate that was used to normalize the band intensity among experiments. (B) Diagram demonstrating the profile of ERK1/2 activation (levels of pERK1/2). The data for the diagram were collected by investigating the level of pERK1/2 in several different clonal lines of HD YAC128 (n=5) and WT iPSCs (n=6). ERK1/2 activation was dramatically decreased in the HD YAC128 iPSCs 10 min after stimulation using bFGF, whereas ERK1/2 activation in the WT iPSCs remained high. Two-way ANOVA; time×genotype interaction, P=0.0013; Bonferroni post-test. (C) Western blot analysis of the levels of total ERK1/2 proteins. (D) The analysis revealed a decreased level of total ERK1 in the YAC128 lines (t-test, P=0.0189). * indicates the YAC128/Oct-eGFP lines.
Mentions: The ERK pathway has been implicated in HD pathogenesis (Bowles and Jones, 2014), and upregulation of its activity might be beneficial in mouse HD models (Maher et al., 2011). Therefore, we investigated the profile of ERK activation in YAC128 iPSCs expressing human mutant huntingtin and investigated whether HD cells in an ESC-like state exhibited dysregulation of the ERK pathway. We stimulated both YAC128 and WT iPSCs using bFGF for 5, 10 or 15 min and compared their levels of ERK pathway activation (Fig. 3A). We observed different profiles of induction of this pathway for WT and HD YAC128 iPSCs (time×genotype interaction, P=0.0013; main effect of time, P<0.0001; two-way ANOVA). In WT iPSCs, we observed a maximum of approximately a 29-fold increase in the ERK1/2 phosphorylation at 5 min compared with basal levels, whereas the level of activation observed at 15 min after stimulation had decreased to 11-fold above basal levels. In YAC128 iPSCs, we observed a similar peak of ERK1/2 activation at 5 min after stimulation. However, the activation of ERK1/2 recorded at 10 min after stimulation had decreased 2.3-fold, whereas the level of ERK1/2 activation in WT iPSCs remained unchanged (P<0.01, Bonferroni post-test; Fig. 3B). In addition, we found that the level of basal activation of ERK1/2 in YAC128 iPSCs was 1.56-fold higher than that in WT iPSCs, although this difference was not significant (P=0.08). Moreover, the analyses of the levels of the total ERK1 and 2 proteins revealed that the total ERK1 protein levels were significantly lower in the YAC128-iPSC cells (Fig. 3C,D).Fig. 3.

Bottom Line: Several molecular and developmental effects of HD have been identified using neural stem cells (NSCs) and differentiated cells, such as neurons and astrocytes.Surprisingly, we found that a number of changes affecting cellular processes in HD were also present in undifferentiated pluripotent HD iPSCs, including the dysregulation of the MAPK and Wnt signaling pathways and the dysregulation of the expression of genes related to oxidative stress, such as Sod1.Interestingly, a common protein interactor of the huntingtin protein and the proteins in the above pathways is p53, and the expression of p53 was dysregulated in HD YAC128 iPSCs and human HD iPSCs.

View Article: PubMed Central - PubMed

Affiliation: Institute of Bioorganic Chemistry, Polish Academy of Sciences, Noskowskiego 12/14, Poznań 61-704, Poland.

No MeSH data available.


Related in: MedlinePlus