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Transcriptome analysis of synaptoneurosomes identifies neuroplasticity genes overexpressed in incipient Alzheimer's disease.

Williams C, Mehrian Shai R, Wu Y, Hsu YH, Sitzer T, Spann B, McCleary C, Mo Y, Miller CA - PLoS ONE (2009)

Bottom Line: These patients also showed increased expression of neuroplasticity related genes, many encoding 3'UTR consensus sequences that regulate translation in the synapse.An increase in mRNA encoding the GluR2 subunit of the alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptor (AMPAR) was paralleled by elevated expression of the corresponding protein in IAD.These results imply a functional impact on synaptic transmission as GluR2, if inserted, maintains the receptors in a low conductance state.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Keck School of Medicine University of Southern California, Los Angeles, California, United States of America.

ABSTRACT
In Alzheimer's disease (AD), early deficits in learning and memory are a consequence of synaptic modification induced by toxic beta-amyloid oligomers (oAbeta). To identify immediate molecular targets downstream of oAbeta binding, we prepared synaptoneurosomes from prefrontal cortex of control and incipient AD (IAD) patients, and isolated mRNAs for comparison of gene expression. This novel approach concentrates synaptic mRNA, thereby increasing the ratio of synaptic to somal mRNA and allowing discrimination of expression changes in synaptically localized genes. In IAD patients, global measures of cognition declined with increasing levels of dimeric Abeta (dAbeta). These patients also showed increased expression of neuroplasticity related genes, many encoding 3'UTR consensus sequences that regulate translation in the synapse. An increase in mRNA encoding the GluR2 subunit of the alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptor (AMPAR) was paralleled by elevated expression of the corresponding protein in IAD. These results imply a functional impact on synaptic transmission as GluR2, if inserted, maintains the receptors in a low conductance state. Some overexpressed genes may induce early deficits in cognition and others compensatory mechanisms, providing targets for intervention to moderate the response to dAbeta.

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GluR2 protein expression.To determine if increased GluR2 mRNA generates elevated protein expression, we compared subjects with a range of MMSE values. Representative immunoblots of A) homogenates and B) synaptoneurosomes, (10 µg protein per lane) were detected with antibodies for GluR2 and GAPDH. C) In controls, the increased ratio (2-fold) of synaptoneurosome (white bar) GluR2 to total GluR2 in homogenates (* p<0.0002) is an indication of synaptic protein enrichment and this ratio increases by 50% in IAD patients (p<5.8E-08). In the IAD group there is a significant increase in synaptoneurosome GluR2 protein expression compared to control (** p<6.0E-05). Homogenate GluR2 (black bar) remains constant in controls and IAD. Data are mean values ±SEM for 4 controls and 4 IAD analyzed in 5 separate experiments. The ratio of GluR2 to GAPDH is given in arbitrary units based on the integrated density value which is the sum of all pixel values after background correction (IDV).
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pone-0004936-g005: GluR2 protein expression.To determine if increased GluR2 mRNA generates elevated protein expression, we compared subjects with a range of MMSE values. Representative immunoblots of A) homogenates and B) synaptoneurosomes, (10 µg protein per lane) were detected with antibodies for GluR2 and GAPDH. C) In controls, the increased ratio (2-fold) of synaptoneurosome (white bar) GluR2 to total GluR2 in homogenates (* p<0.0002) is an indication of synaptic protein enrichment and this ratio increases by 50% in IAD patients (p<5.8E-08). In the IAD group there is a significant increase in synaptoneurosome GluR2 protein expression compared to control (** p<6.0E-05). Homogenate GluR2 (black bar) remains constant in controls and IAD. Data are mean values ±SEM for 4 controls and 4 IAD analyzed in 5 separate experiments. The ratio of GluR2 to GAPDH is given in arbitrary units based on the integrated density value which is the sum of all pixel values after background correction (IDV).

Mentions: GluR2 was selected to determine if protein expression correlates with the increased mRNA seen in synaptoneurosomes from individual patients. Equal amounts of protein (25 µg) from homogenate and synaptoneurosome preparations were separated by polyacrylamide gel electrophoresis (PAGE). Immunoblots labeled with anti-GluR2 and GAPDH revealed that, in controls, there is a significant increase in the GluR2 subunit in synaptoneurosomes compared to whole homogenates (p<0.0002), confirming the data presented in Figure 1. In synaptoneurosomes from IAD patients, GluR2 increased as MMSE declined, whereas total GluR2 in homogenates remained relatively stable (Figure 5A, B, C). In Figure 5C, the ratio of synaptoneurosome-associated GluR2 to total GluR2 present in homogenates increases 2-fold in controls to more than 3-fold in IAD. The increase in synaptic GluR2 subunits is masked in homogenates. Thus, IAD patients have increased levels of GluR2 mRNA at the synapse which generate an increase in receptor subunits.


Transcriptome analysis of synaptoneurosomes identifies neuroplasticity genes overexpressed in incipient Alzheimer's disease.

Williams C, Mehrian Shai R, Wu Y, Hsu YH, Sitzer T, Spann B, McCleary C, Mo Y, Miller CA - PLoS ONE (2009)

GluR2 protein expression.To determine if increased GluR2 mRNA generates elevated protein expression, we compared subjects with a range of MMSE values. Representative immunoblots of A) homogenates and B) synaptoneurosomes, (10 µg protein per lane) were detected with antibodies for GluR2 and GAPDH. C) In controls, the increased ratio (2-fold) of synaptoneurosome (white bar) GluR2 to total GluR2 in homogenates (* p<0.0002) is an indication of synaptic protein enrichment and this ratio increases by 50% in IAD patients (p<5.8E-08). In the IAD group there is a significant increase in synaptoneurosome GluR2 protein expression compared to control (** p<6.0E-05). Homogenate GluR2 (black bar) remains constant in controls and IAD. Data are mean values ±SEM for 4 controls and 4 IAD analyzed in 5 separate experiments. The ratio of GluR2 to GAPDH is given in arbitrary units based on the integrated density value which is the sum of all pixel values after background correction (IDV).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2654156&req=5

pone-0004936-g005: GluR2 protein expression.To determine if increased GluR2 mRNA generates elevated protein expression, we compared subjects with a range of MMSE values. Representative immunoblots of A) homogenates and B) synaptoneurosomes, (10 µg protein per lane) were detected with antibodies for GluR2 and GAPDH. C) In controls, the increased ratio (2-fold) of synaptoneurosome (white bar) GluR2 to total GluR2 in homogenates (* p<0.0002) is an indication of synaptic protein enrichment and this ratio increases by 50% in IAD patients (p<5.8E-08). In the IAD group there is a significant increase in synaptoneurosome GluR2 protein expression compared to control (** p<6.0E-05). Homogenate GluR2 (black bar) remains constant in controls and IAD. Data are mean values ±SEM for 4 controls and 4 IAD analyzed in 5 separate experiments. The ratio of GluR2 to GAPDH is given in arbitrary units based on the integrated density value which is the sum of all pixel values after background correction (IDV).
Mentions: GluR2 was selected to determine if protein expression correlates with the increased mRNA seen in synaptoneurosomes from individual patients. Equal amounts of protein (25 µg) from homogenate and synaptoneurosome preparations were separated by polyacrylamide gel electrophoresis (PAGE). Immunoblots labeled with anti-GluR2 and GAPDH revealed that, in controls, there is a significant increase in the GluR2 subunit in synaptoneurosomes compared to whole homogenates (p<0.0002), confirming the data presented in Figure 1. In synaptoneurosomes from IAD patients, GluR2 increased as MMSE declined, whereas total GluR2 in homogenates remained relatively stable (Figure 5A, B, C). In Figure 5C, the ratio of synaptoneurosome-associated GluR2 to total GluR2 present in homogenates increases 2-fold in controls to more than 3-fold in IAD. The increase in synaptic GluR2 subunits is masked in homogenates. Thus, IAD patients have increased levels of GluR2 mRNA at the synapse which generate an increase in receptor subunits.

Bottom Line: These patients also showed increased expression of neuroplasticity related genes, many encoding 3'UTR consensus sequences that regulate translation in the synapse.An increase in mRNA encoding the GluR2 subunit of the alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptor (AMPAR) was paralleled by elevated expression of the corresponding protein in IAD.These results imply a functional impact on synaptic transmission as GluR2, if inserted, maintains the receptors in a low conductance state.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Keck School of Medicine University of Southern California, Los Angeles, California, United States of America.

ABSTRACT
In Alzheimer's disease (AD), early deficits in learning and memory are a consequence of synaptic modification induced by toxic beta-amyloid oligomers (oAbeta). To identify immediate molecular targets downstream of oAbeta binding, we prepared synaptoneurosomes from prefrontal cortex of control and incipient AD (IAD) patients, and isolated mRNAs for comparison of gene expression. This novel approach concentrates synaptic mRNA, thereby increasing the ratio of synaptic to somal mRNA and allowing discrimination of expression changes in synaptically localized genes. In IAD patients, global measures of cognition declined with increasing levels of dimeric Abeta (dAbeta). These patients also showed increased expression of neuroplasticity related genes, many encoding 3'UTR consensus sequences that regulate translation in the synapse. An increase in mRNA encoding the GluR2 subunit of the alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptor (AMPAR) was paralleled by elevated expression of the corresponding protein in IAD. These results imply a functional impact on synaptic transmission as GluR2, if inserted, maintains the receptors in a low conductance state. Some overexpressed genes may induce early deficits in cognition and others compensatory mechanisms, providing targets for intervention to moderate the response to dAbeta.

Show MeSH
Related in: MedlinePlus