Altered regulation of tau phosphorylation in a mouse model of down syndrome aging - PubMed (original) (raw)

Altered regulation of tau phosphorylation in a mouse model of down syndrome aging

Olivia Sheppard et al. Neurobiol Aging. 2012 Apr.

Abstract

Down syndrome (DS) results from trisomy of human chromosome 21 (Hsa21) and is associated with an increased risk of Alzheimer's disease (AD). Here, using the unique transchromosomic Tc1 mouse model of DS we investigate the influence of trisomy of Hsa21 on the protein tau, which is hyperphosphorylated in Alzheimer's disease. We show that in old, but not young, Tc1 mice increased phosphorylation of tau occurs at a site suggested to be targeted by the Hsa21 encoded kinase, dual-specificity tyrosine-(Y)-phosphorylation regulated kinase 1A (DYRK1A). We show that DYRK1A is upregulated in young and old Tc1 mice, but that young trisomic mice may be protected from accumulating aberrantly phosphorylated tau. We observe that the key tau kinase, glycogen synthase kinase3-β (GSK-3β) is aberrantly phosphorylated at an inhibitory site in the aged Tc1 brain which may reduce total glycogen synthase kinase3-β activity. It is possible that a similar mechanism may also occur in people with DS.

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Figures

Fig. 1

Tau is aberrantly phosphorylated at threonine 212 in the hippocampus of aged but not young Tc1 mice. The abundance of phosphorylated forms of tau was investigated by Western blot of total hippocampal protein lysates using anti-phospho-tau antibodies Thr212 (A) and (D), AT8 (B) and (E), and PHF1 (C) and (F). Equal amounts of total protein were loaded per lane and anti-total tau and β-actin antibodies were used as controls (A–F). Relative phospho-tau/total-tau signal was determine for each Tc1 sample and compared with the phospho-tau/total-tau ratio detected in the matched littermate nontranschromosomic control, in both young (2 months of age) and old (20 months of age) animals. Normalized signals were analyzed by analysis of variance (ANOVA) including the fixed factor of tissue type (hippocampus or cortex) and no significant effect of tissue was observed therefore data for both types of tissues was combined for subsequent analysis. A further ANOVA using fixed factors genotype of mouse (Tc1 or control) and age of mouse (2 months of age or 20 months of age) was performed. Tau Thr212 (normalized to total tau) signal was significantly affected by the interaction of genotype of mouse × age (F(1,51) = 6.110; p = 0.17). Post hoc tests showed that a significant increase in tau phosphorylated at Thr212 was detected in old Tc1 hippocampus compared with age- and sex-matched nontranschromosomic littermate control (least significant difference [LSD] p = 0.017; n = 8) and also old Tc1 hippocampus compared with young Tc1 hippocampus (LSD p = 0.04; n = 7). No significant increase in tau phosphorylated at Thr212 was detected in the hippocampus of young Tc1 mice compared with age- and sex-matched controls (2 months of age) (n = 7). No significant increase in phospho-tau detected by AT8 or PHF1 was detected in aged or young Tc1 hippocampus compared with wild type littermate control hippocampus (n = 9–10). Error bars show standard error of the mean.

Fig. 2

Tau is aberrantly phosphorylated at threonine 212 in the cortex of aged Tc1 mice. The abundance of phosphorylated forms of tau was investigated by Western blot of total cortical protein lysates using anti-phospho-tau antibodies Thr212 (A), AT8 (B), and PHF1 (C). Equal amounts of total protein were loaded per lane and anti-total tau and β-actin antibodies were used as controls (A–C). Relative phospho-tau/total-tau signal was determine for each Tc1 sample and compared with the phospho-tau/total-tau ratio detected in the matched littermate nontranschromosomic control. Post hoc tests showed that a significant increase in tau phosphorylated at Thr212 was detected in old Tc1 cortex (20 months of age) compared with matched wild type littermate control cortex (LSD p < 0.04; n = 10). No significant increase in phospho-tau detected by AT8 or PHF1 was detected in aged or young Tc1 cortex compared with euploid control cortex (n = 7–10). Error bars show standard error of the mean.

Fig. 3

The expression of DYRK1A is elevated in the brains of Tc1 mice compared with wild type littermate control mice. The expression of mouse Dyrk1a and human DYRK1A transcript in Tc1 mouse brain was confirmed by RT-polymerase chain reaction (PCR) on total brain RNA from young Tc1 mice (2 months of age, n = 3). Primers that cross-react with both human and mouse Dyrk1A transcript product size 235 base pairs (A and B) were multiplexed with either specific for mouse Dyrk1a that produce a 793-base pair product (A) or primers specific for human DYRK1A that produce a 621-base pair product (B). The amount of DYRK1A protein in the hippocampus and cortex of Tc1 mice was quantified by Western blot of total protein lysates using anti-DYRK1A antibody 7D11, which is predicted to recognize both the human and mouse form of the protein (D–F). Equal amounts of total protein were loaded per lane and anti-β-actin and GAPDH antibodies were used as controls for total protein amount per lane (F) and (G). Relative DYRK1A/β-actin signal was determined for each Tc1 sample and compared with the signal for the matched wild type littermate control. An increase in DYRK1A was detected in the hippocampus and cortex of young (2 months of age) and old (20 months of age) Tc1 mice compared with wild type littermate control mice (analysis of variance [ANOVA] genotype F(1,47) = 723.076; p = 0.000; post hoc least significant difference [LSD] young hippocampus, p = 0.028, young cortex, p = 0.004, old hippocampus, p = 0.041, and old cortex, p = 0.000). Error bars show standard error of the mean.

Fig. 4

Phosphorylation of glycogen synthase kinase3-β (GSK-3β) at serine-9 is increased in the hippocampus and cortex of old Tc1 mice. The abundance of total GSK-3β and GSK-3β phosphorylated at Ser9 and Tyr216 was investigated by Western blot of total hippocampal (A–C) and (G–I) and cortical (D–F) protein lysates, from old (A–F) and young (G–I) Tc1 mice and matched wild type littermate control animals. Equal amounts of total protein were loaded per lane and anti-β-actin antibody signal was used as a control for amount of total protein loaded (A–I). Total GSK-3β signal was normalized to GAPDH signal in Tc1 samples, and compared with age- and sex-matched nontranschromosomic control signal. No significant difference in total GSK-3β in Tc1 samples was detected in old (20 months of age) hippocampus (n = 10) (A), old (20 months of age) cortex (n = 7) (D), or young (2 months of age) hippocampus (n = 7) (G). Phospho-GSK-3β signal was normalized to total GSK-3β signal in Tc1 samples, and compared with the respective matched littermate euploid control signal. No significant difference in GSK-3β phosphorylated at Tyr216 was detected in Tc1 compared with controls in old (20 months of age) hippocampus (n = 10) (B), old (20 months of age) cortex (n = 6) (E), or young (2 months of age) hippocampus (n = 6) (H). A significant increase GSK-3β phosphorylated at Ser9 was detected in old (20 months) Tc1 brain (analysis of variance [ANOVA] genotype × age F(1,45) = 5.482; p = 0.024) by post hoc least significant difference (LSD) tests significant increases in signal were detected in aged Tc1 hippocampus (p = 0.011) (C) and old (20 months of age) cortex (p = 0.003) (F) compared with age- and sex-matched control samples. No significant increase in phosphorylation of GSK-3β at Ser9 was detected in young Tc1 hippocampus (n = 7) (I). Error bars show standard error of the mean.

Fig. 5

Phosphorylation of AKT at serine-473 is increased in the cortex of old Tc1 mice. The abundance of total AKT and AKT phosphorylated at Ser473 was investigated by Western blot of total hippocampal (A–B) and (E–G) and cortical (C–D) and (H) protein lysates, from young (2 months of age) (E) and (F) and old (20 months of age) (A–D) and (G–H) Tc1 mice and age- and sex-matched wild type littermate control animals. Equal amounts of total protein were loaded per lane and anti-β-actin antibody signal was used as a control for amount of total protein loaded (A–H). Total AKT signal was normalized to β-actin signal in Tc1 samples, and compared with the respective matched wild type littermate control signal. No significant difference in total AKT in Tc1 samples was detected in old (20 months of age) hippocampus (n = 12) (A) or cortex (n = 12) (C). A significant increase in total AKT signal was detected in young hippocampus (analysis of variance [ANOVA] genotype F(1,47) = 4.570; p = 0.038; least significant difference [LSD] post hoc p = 0.024) (E). Phospho-AKT signal was normalized to total AKT signal in Tc1 samples, and compared with the respective matched littermate control signal. A significant increase in AKT phosphorylated at Ser473 was detected in old (20 months) Tc1 cortex (ANOVA genotype F(1,54) = 4.975; p = 0.030; post hoc LSD p = 0.013) (D). Error bars show standard error of the mean.

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Altered regulation of tau phosphorylation in a mouse model of down syndrome aging - PubMed (original) (raw)