Limits...
In Absence of the Cellular Prion Protein, Alterations in Copper Metabolism and Copper-Dependent Oxidase Activity Affect Iron Distribution

View Article: PubMed Central - PubMed

ABSTRACT

Essential elements as copper and iron modulate a wide range of physiological functions. Their metabolism is strictly regulated by cellular pathways, since dysregulation of metal homeostasis is responsible for many detrimental effects. Neurodegenerative disorders such as Alzheimer's disease, Parkinson's disease and prion diseases are characterized by alterations of metal ions. These neurodegenerative maladies involve proteins that bind metals and mediate their metabolism through not well-defined mechanisms. Prion protein, for instance, interacts with divalent cations via multiple metal-binding sites and it modulates several metal-dependent physiological functions, such as S-nitrosylation of NMDA receptors. In this work we focused on the effect of prion protein absence on copper and iron metabolism during development and adulthood. In particular, we investigated copper and iron functional values in serum and several organs such as liver, spleen, total brain and isolated hippocampus. Our results show that iron content is diminished in prion protein- mouse serum, while it accumulates in liver and spleen. Our data suggest that these alterations can be due to impairments in copper-dependent cerulopalsmin activity which is known to affect iron mobilization. In prion protein- mouse total brain and hippocampus, metal ion content shows a fluctuating trend, suggesting the presence of homeostatic compensatory mechanisms. However, copper and iron functional values are likely altered also in these two organs, as indicated by the modulation of metal-binding protein expression levels. Altogether, these results reveal that the absence of the cellular prion protein impairs copper metabolism and copper-dependent oxidase activity, with ensuing alteration of iron mobilization from cellular storage compartments.

No MeSH data available.


Analysis of Cu and Fe content and metal-binding protein expression in wild-type and PrPC- mouse isolated hippocampus at different ages. (A) The graph shows the ratio of Cu and Fe levels in Prnp0/0 and Prnp+/+ hippocampus samples (P1, P180, P365 N = 4; P7 N = 7; P30 N = 6; P90 N = 5). (B) Representative Western blot images showing metal-binding protein levels in Prnp0/0 and Prnp+/+ hippocampal samples (N = 4). The constant level of the housekeeping proteins (β-III Tubulin and β-Actin) are also reported. (C) The graph shows the up- or down-regulation of protein expression in Prnp0/0 samples compared to Prnp+/+, i.e., (Prnp0/0 protein OD/housekeeping OD)/(Prnp+/+ protein OD/housekeeping OD). All error bars indicate SD; N = 4 minimum); *p < 0.05; **p < 0.01; ***p < 0.001.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC5037227&req=5

Figure 5: Analysis of Cu and Fe content and metal-binding protein expression in wild-type and PrPC- mouse isolated hippocampus at different ages. (A) The graph shows the ratio of Cu and Fe levels in Prnp0/0 and Prnp+/+ hippocampus samples (P1, P180, P365 N = 4; P7 N = 7; P30 N = 6; P90 N = 5). (B) Representative Western blot images showing metal-binding protein levels in Prnp0/0 and Prnp+/+ hippocampal samples (N = 4). The constant level of the housekeeping proteins (β-III Tubulin and β-Actin) are also reported. (C) The graph shows the up- or down-regulation of protein expression in Prnp0/0 samples compared to Prnp+/+, i.e., (Prnp0/0 protein OD/housekeeping OD)/(Prnp+/+ protein OD/housekeeping OD). All error bars indicate SD; N = 4 minimum); *p < 0.05; **p < 0.01; ***p < 0.001.

Mentions: Since CNS is the organ with the highest PrPC expression (Figure S3) and primary target of prion disorders, we analyzed the impact of PrPC absence on copper and iron metabolism. We analyzed the total brain and the isolated hippocampus. The latter is the region showing PrPC expression at synapse and the most prominent alterations in Prnp0/0 mouse model. We first measured copper and iron content in wild-type and PrPC- mouse total brain and isolated hippocampus at different ages, from P1 to 1-year-old, and expressed results as the ratio between Prnp0/0 and Prnp+/+ ion concentration values (Figures 4A, 5A). Results expressed in μg/mL are reported in Figures S1H–K. Both copper and iron show a fluctuating behavior along ages in both total brain and hippocampus, with a reduction in their content at early stages (P1 and P7) and adulthood (P180), and an increase at P30-P90 (Figures 4A, 5A).


In Absence of the Cellular Prion Protein, Alterations in Copper Metabolism and Copper-Dependent Oxidase Activity Affect Iron Distribution
Analysis of Cu and Fe content and metal-binding protein expression in wild-type and PrPC- mouse isolated hippocampus at different ages. (A) The graph shows the ratio of Cu and Fe levels in Prnp0/0 and Prnp+/+ hippocampus samples (P1, P180, P365 N = 4; P7 N = 7; P30 N = 6; P90 N = 5). (B) Representative Western blot images showing metal-binding protein levels in Prnp0/0 and Prnp+/+ hippocampal samples (N = 4). The constant level of the housekeeping proteins (β-III Tubulin and β-Actin) are also reported. (C) The graph shows the up- or down-regulation of protein expression in Prnp0/0 samples compared to Prnp+/+, i.e., (Prnp0/0 protein OD/housekeeping OD)/(Prnp+/+ protein OD/housekeeping OD). All error bars indicate SD; N = 4 minimum); *p < 0.05; **p < 0.01; ***p < 0.001.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5037227&req=5

Figure 5: Analysis of Cu and Fe content and metal-binding protein expression in wild-type and PrPC- mouse isolated hippocampus at different ages. (A) The graph shows the ratio of Cu and Fe levels in Prnp0/0 and Prnp+/+ hippocampus samples (P1, P180, P365 N = 4; P7 N = 7; P30 N = 6; P90 N = 5). (B) Representative Western blot images showing metal-binding protein levels in Prnp0/0 and Prnp+/+ hippocampal samples (N = 4). The constant level of the housekeeping proteins (β-III Tubulin and β-Actin) are also reported. (C) The graph shows the up- or down-regulation of protein expression in Prnp0/0 samples compared to Prnp+/+, i.e., (Prnp0/0 protein OD/housekeeping OD)/(Prnp+/+ protein OD/housekeeping OD). All error bars indicate SD; N = 4 minimum); *p < 0.05; **p < 0.01; ***p < 0.001.
Mentions: Since CNS is the organ with the highest PrPC expression (Figure S3) and primary target of prion disorders, we analyzed the impact of PrPC absence on copper and iron metabolism. We analyzed the total brain and the isolated hippocampus. The latter is the region showing PrPC expression at synapse and the most prominent alterations in Prnp0/0 mouse model. We first measured copper and iron content in wild-type and PrPC- mouse total brain and isolated hippocampus at different ages, from P1 to 1-year-old, and expressed results as the ratio between Prnp0/0 and Prnp+/+ ion concentration values (Figures 4A, 5A). Results expressed in μg/mL are reported in Figures S1H–K. Both copper and iron show a fluctuating behavior along ages in both total brain and hippocampus, with a reduction in their content at early stages (P1 and P7) and adulthood (P180), and an increase at P30-P90 (Figures 4A, 5A).

View Article: PubMed Central - PubMed

ABSTRACT

Essential elements as copper and iron modulate a wide range of physiological functions. Their metabolism is strictly regulated by cellular pathways, since dysregulation of metal homeostasis is responsible for many detrimental effects. Neurodegenerative disorders such as Alzheimer's disease, Parkinson's disease and prion diseases are characterized by alterations of metal ions. These neurodegenerative maladies involve proteins that bind metals and mediate their metabolism through not well-defined mechanisms. Prion protein, for instance, interacts with divalent cations via multiple metal-binding sites and it modulates several metal-dependent physiological functions, such as S-nitrosylation of NMDA receptors. In this work we focused on the effect of prion protein absence on copper and iron metabolism during development and adulthood. In particular, we investigated copper and iron functional values in serum and several organs such as liver, spleen, total brain and isolated hippocampus. Our results show that iron content is diminished in prion protein- mouse serum, while it accumulates in liver and spleen. Our data suggest that these alterations can be due to impairments in copper-dependent cerulopalsmin activity which is known to affect iron mobilization. In prion protein- mouse total brain and hippocampus, metal ion content shows a fluctuating trend, suggesting the presence of homeostatic compensatory mechanisms. However, copper and iron functional values are likely altered also in these two organs, as indicated by the modulation of metal-binding protein expression levels. Altogether, these results reveal that the absence of the cellular prion protein impairs copper metabolism and copper-dependent oxidase activity, with ensuing alteration of iron mobilization from cellular storage compartments.

No MeSH data available.