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De-regulation of gene expression and alternative splicing affects distinct cellular pathways in the aging hippocampus.

Stilling RM, Benito E, Gertig M, Barth J, Capece V, Burkhardt S, Bonn S, Fischer A - Front Cell Neurosci (2014)

Bottom Line: This approach enabled us to test differential expression of coding and non-coding transcripts, as well as differential splicing and RNA editing.We report a specific age-associated gene expression signature that is associated with major genetic risk factors for late-onset AD (LOAD).This signature is dominated by neuroinflammatory processes, specifically activation of the complement system at the level of increased gene expression, while de-regulation of neuronal plasticity appears to be mediated by compromised RNA splicing.

View Article: PubMed Central - PubMed

Affiliation: Department of Psychiatry and Psychotherapy, University Medical Center Göttingen Göttingen, Germany ; Research Group for Epigenetics in Neurodegenerative Diseases, German Center for Neurodegenerative Diseases (DZNE) Göttingen Göttingen, Germany.

ABSTRACT
Aging is accompanied by gradually increasing impairment of cognitive abilities and constitutes the main risk factor of neurodegenerative conditions like Alzheimer's disease (AD). The underlying mechanisms are however not well understood. Here we analyze the hippocampal transcriptome of young adult mice and two groups of mice at advanced age using RNA sequencing. This approach enabled us to test differential expression of coding and non-coding transcripts, as well as differential splicing and RNA editing. We report a specific age-associated gene expression signature that is associated with major genetic risk factors for late-onset AD (LOAD). This signature is dominated by neuroinflammatory processes, specifically activation of the complement system at the level of increased gene expression, while de-regulation of neuronal plasticity appears to be mediated by compromised RNA splicing.

No MeSH data available.


Related in: MedlinePlus

Aging is associated with changes in gene expression and upregulation of immune system functions in the hippocampus. (A) Heatmaps of differentially expressed genes in 3-month (3M) and 24-month-old mice (24M, left panel) and 3 and 29-month (29M) old mice (right panel). High expression is marked by the red color spectrum, low expression by blue colors. (B) Pie chart showing the percentage of genes up-and down-regulated in 3 vs. 24/29 month-old mice. (C) Venn diagrams comparing significantly regulated genes found in the different experiments (3M vs. 24M and 3M vs. 29M). (D) Genes up-regulated at 24 and 29 month of age were subjected to functional enrichment analysis for overrepresented biological processes and KEGG pathways. (E) Fold enrichment of overrepresented KEGG pathways in up-regulated genes when individually comparing the 3M vs. 24M and 3M vs. 29M groups. (F) Using non-cutoff gene set enrichment analysis (GSEA), the top up-regulated KEGG pathway was the complement and coagulation cascade. Shown are enrichment plots along expressed genes, ranked by fold-change [left/red: positive log2-fold-change (upregulation), right/blue: negative log2-fold-change (downregulation); a gene contributing to the enrichment score of the selected pathway along the ranked list is marked by a vertical black line and increases the cumulative enrichment score (green line)]. (G) qPCR analysis of the C4b gene (***p < 0.001 vs. 3M group, two-sided t-tests; #p < 0.05 vs. 24M group, two-sided t-test; n = 5[3M]/4[24M]/8[29M]). Error bars indicate s.e.m.
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Figure 2: Aging is associated with changes in gene expression and upregulation of immune system functions in the hippocampus. (A) Heatmaps of differentially expressed genes in 3-month (3M) and 24-month-old mice (24M, left panel) and 3 and 29-month (29M) old mice (right panel). High expression is marked by the red color spectrum, low expression by blue colors. (B) Pie chart showing the percentage of genes up-and down-regulated in 3 vs. 24/29 month-old mice. (C) Venn diagrams comparing significantly regulated genes found in the different experiments (3M vs. 24M and 3M vs. 29M). (D) Genes up-regulated at 24 and 29 month of age were subjected to functional enrichment analysis for overrepresented biological processes and KEGG pathways. (E) Fold enrichment of overrepresented KEGG pathways in up-regulated genes when individually comparing the 3M vs. 24M and 3M vs. 29M groups. (F) Using non-cutoff gene set enrichment analysis (GSEA), the top up-regulated KEGG pathway was the complement and coagulation cascade. Shown are enrichment plots along expressed genes, ranked by fold-change [left/red: positive log2-fold-change (upregulation), right/blue: negative log2-fold-change (downregulation); a gene contributing to the enrichment score of the selected pathway along the ranked list is marked by a vertical black line and increases the cumulative enrichment score (green line)]. (G) qPCR analysis of the C4b gene (***p < 0.001 vs. 3M group, two-sided t-tests; #p < 0.05 vs. 24M group, two-sided t-test; n = 5[3M]/4[24M]/8[29M]). Error bars indicate s.e.m.

Mentions: In the next step, we isolated total RNA from the hippocampi of 3-, 24-, and 29-month-old mice and subjected it to RNA sequencing. In line with previous studies, we observed that aging was not associated with massive changes in cell number (Long et al., 1999) (Figure S1). We first compared differential gene expression across the different age groups using the 3-month group as reference. 477 genes were differentially expressed (313 up-regulated, 164 down-regulated) in 24-month-old mice and 323 genes (275 up-regulated, 48 down-regulated) in 29-month-old mice (Figure 2A, Table S1) when compared to the 3-month group. In all comparisons, we observed a general trend toward higher numbers of up-regulated genes (~70%) compared to down-regulated genes (Figure 2B). When we compared these lists among each other, we found a significant amount of overlap between genes up-regulated at 24 and 29 months of age (122 genes, Figure 2C, Table S1). The overlap between the genes down-regulated in 24- and 29-month-old mice was less pronounced (17 genes, Figure 2C, Table S1). When we analyzed the 122 genes commonly up-regulated in 24- and 29-month-old mice for functional pathways we observed inflammatory signaling pathways, namely the “Systemic lupus erythematosus” and the “complement and coagulation pathway” to be highly enriched (Figure 2D, Table S2). It has to be noted that several genes of the complement system are linked to systemic lupus erythematosus, which explains the enrichment of this pathway (Table S2). This data suggests that activation of the complement system is one of the key features of the aging hippocampus. Similar results were obtained when we separately analyzed all genes up-regulated in 3 vs. 24-month-old mice (Figure 2E, left panel). When we analyzed all up-regulated genes in 3 vs. 29-month-old mice, we found significant enrichment of additional immune-related pathways (Figure 2E, right panel, blue bars), suggestive of an even more pronounced immune activation with increasing age. Another group of up-regulated genes was associated with cell adhesion since “cell adhesion molecules” was also identified as a significantly enriched pathway in 24- and 29-month-old mice (Figure 2E, Table S2).


De-regulation of gene expression and alternative splicing affects distinct cellular pathways in the aging hippocampus.

Stilling RM, Benito E, Gertig M, Barth J, Capece V, Burkhardt S, Bonn S, Fischer A - Front Cell Neurosci (2014)

Aging is associated with changes in gene expression and upregulation of immune system functions in the hippocampus. (A) Heatmaps of differentially expressed genes in 3-month (3M) and 24-month-old mice (24M, left panel) and 3 and 29-month (29M) old mice (right panel). High expression is marked by the red color spectrum, low expression by blue colors. (B) Pie chart showing the percentage of genes up-and down-regulated in 3 vs. 24/29 month-old mice. (C) Venn diagrams comparing significantly regulated genes found in the different experiments (3M vs. 24M and 3M vs. 29M). (D) Genes up-regulated at 24 and 29 month of age were subjected to functional enrichment analysis for overrepresented biological processes and KEGG pathways. (E) Fold enrichment of overrepresented KEGG pathways in up-regulated genes when individually comparing the 3M vs. 24M and 3M vs. 29M groups. (F) Using non-cutoff gene set enrichment analysis (GSEA), the top up-regulated KEGG pathway was the complement and coagulation cascade. Shown are enrichment plots along expressed genes, ranked by fold-change [left/red: positive log2-fold-change (upregulation), right/blue: negative log2-fold-change (downregulation); a gene contributing to the enrichment score of the selected pathway along the ranked list is marked by a vertical black line and increases the cumulative enrichment score (green line)]. (G) qPCR analysis of the C4b gene (***p < 0.001 vs. 3M group, two-sided t-tests; #p < 0.05 vs. 24M group, two-sided t-test; n = 5[3M]/4[24M]/8[29M]). Error bars indicate s.e.m.
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Figure 2: Aging is associated with changes in gene expression and upregulation of immune system functions in the hippocampus. (A) Heatmaps of differentially expressed genes in 3-month (3M) and 24-month-old mice (24M, left panel) and 3 and 29-month (29M) old mice (right panel). High expression is marked by the red color spectrum, low expression by blue colors. (B) Pie chart showing the percentage of genes up-and down-regulated in 3 vs. 24/29 month-old mice. (C) Venn diagrams comparing significantly regulated genes found in the different experiments (3M vs. 24M and 3M vs. 29M). (D) Genes up-regulated at 24 and 29 month of age were subjected to functional enrichment analysis for overrepresented biological processes and KEGG pathways. (E) Fold enrichment of overrepresented KEGG pathways in up-regulated genes when individually comparing the 3M vs. 24M and 3M vs. 29M groups. (F) Using non-cutoff gene set enrichment analysis (GSEA), the top up-regulated KEGG pathway was the complement and coagulation cascade. Shown are enrichment plots along expressed genes, ranked by fold-change [left/red: positive log2-fold-change (upregulation), right/blue: negative log2-fold-change (downregulation); a gene contributing to the enrichment score of the selected pathway along the ranked list is marked by a vertical black line and increases the cumulative enrichment score (green line)]. (G) qPCR analysis of the C4b gene (***p < 0.001 vs. 3M group, two-sided t-tests; #p < 0.05 vs. 24M group, two-sided t-test; n = 5[3M]/4[24M]/8[29M]). Error bars indicate s.e.m.
Mentions: In the next step, we isolated total RNA from the hippocampi of 3-, 24-, and 29-month-old mice and subjected it to RNA sequencing. In line with previous studies, we observed that aging was not associated with massive changes in cell number (Long et al., 1999) (Figure S1). We first compared differential gene expression across the different age groups using the 3-month group as reference. 477 genes were differentially expressed (313 up-regulated, 164 down-regulated) in 24-month-old mice and 323 genes (275 up-regulated, 48 down-regulated) in 29-month-old mice (Figure 2A, Table S1) when compared to the 3-month group. In all comparisons, we observed a general trend toward higher numbers of up-regulated genes (~70%) compared to down-regulated genes (Figure 2B). When we compared these lists among each other, we found a significant amount of overlap between genes up-regulated at 24 and 29 months of age (122 genes, Figure 2C, Table S1). The overlap between the genes down-regulated in 24- and 29-month-old mice was less pronounced (17 genes, Figure 2C, Table S1). When we analyzed the 122 genes commonly up-regulated in 24- and 29-month-old mice for functional pathways we observed inflammatory signaling pathways, namely the “Systemic lupus erythematosus” and the “complement and coagulation pathway” to be highly enriched (Figure 2D, Table S2). It has to be noted that several genes of the complement system are linked to systemic lupus erythematosus, which explains the enrichment of this pathway (Table S2). This data suggests that activation of the complement system is one of the key features of the aging hippocampus. Similar results were obtained when we separately analyzed all genes up-regulated in 3 vs. 24-month-old mice (Figure 2E, left panel). When we analyzed all up-regulated genes in 3 vs. 29-month-old mice, we found significant enrichment of additional immune-related pathways (Figure 2E, right panel, blue bars), suggestive of an even more pronounced immune activation with increasing age. Another group of up-regulated genes was associated with cell adhesion since “cell adhesion molecules” was also identified as a significantly enriched pathway in 24- and 29-month-old mice (Figure 2E, Table S2).

Bottom Line: This approach enabled us to test differential expression of coding and non-coding transcripts, as well as differential splicing and RNA editing.We report a specific age-associated gene expression signature that is associated with major genetic risk factors for late-onset AD (LOAD).This signature is dominated by neuroinflammatory processes, specifically activation of the complement system at the level of increased gene expression, while de-regulation of neuronal plasticity appears to be mediated by compromised RNA splicing.

View Article: PubMed Central - PubMed

Affiliation: Department of Psychiatry and Psychotherapy, University Medical Center Göttingen Göttingen, Germany ; Research Group for Epigenetics in Neurodegenerative Diseases, German Center for Neurodegenerative Diseases (DZNE) Göttingen Göttingen, Germany.

ABSTRACT
Aging is accompanied by gradually increasing impairment of cognitive abilities and constitutes the main risk factor of neurodegenerative conditions like Alzheimer's disease (AD). The underlying mechanisms are however not well understood. Here we analyze the hippocampal transcriptome of young adult mice and two groups of mice at advanced age using RNA sequencing. This approach enabled us to test differential expression of coding and non-coding transcripts, as well as differential splicing and RNA editing. We report a specific age-associated gene expression signature that is associated with major genetic risk factors for late-onset AD (LOAD). This signature is dominated by neuroinflammatory processes, specifically activation of the complement system at the level of increased gene expression, while de-regulation of neuronal plasticity appears to be mediated by compromised RNA splicing.

No MeSH data available.


Related in: MedlinePlus