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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.


Comparison of Cu, Fe, oxidase activity, metal-binding protein expression levels in wild-type and PrPC- mouse liver at different ages. (A) The graph shows the ratio of Cu and Fe levels in Prnp0/0 and Prnp+/+ liver samples (P15, P90 N = 5; P30, P180, P365 N = 4). (B) The graph shows the levels of oxidase activity as U/mL in Prnp0/0 and Prnp+/+ liver (P15, P90, P180, P365 N = 4; P30 N = 5). (C) Representative Western blot images showing metal-binding protein levels in Prnp0/0 and Prnp+/+ liver samples. The constant level of the housekeeping protein (β-Actin) are also reported. (D) 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), N = 4. All error bars indicate SD; *p < 0.05; **p < 0.01.
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Figure 2: Comparison of Cu, Fe, oxidase activity, metal-binding protein expression levels in wild-type and PrPC- mouse liver at different ages. (A) The graph shows the ratio of Cu and Fe levels in Prnp0/0 and Prnp+/+ liver samples (P15, P90 N = 5; P30, P180, P365 N = 4). (B) The graph shows the levels of oxidase activity as U/mL in Prnp0/0 and Prnp+/+ liver (P15, P90, P180, P365 N = 4; P30 N = 5). (C) Representative Western blot images showing metal-binding protein levels in Prnp0/0 and Prnp+/+ liver samples. The constant level of the housekeeping protein (β-Actin) are also reported. (D) 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), N = 4. All error bars indicate SD; *p < 0.05; **p < 0.01.

Mentions: Incorporation of copper into Cp mainly occurs in the liver (Terada et al., 1995). Therefore, we measured copper content in Prnp0/0 and Prnp+/+ mouse liver during early development and adulthood, and expressed results as the ratio between Prnp0/0 and Prnp+/+ ion concentration values (Figure 2A). Results expressed in μg/mL are reported in Figures S1D,E. Similarly to serum, total copper content is not overall altered in Prnp0/0 mouse liver, though a decrease of about 30% occurs at the early stage P15 (Figure 2A). Our data show iron accumulation in the liver starting from P90 (Figure 2A), corresponding to the reduction of serum iron concentration (Figure 1A). As for serum Cp activity, liver oxidase activity is diminished in Prnp0/0 mice (Figure 2B). The activity of ferroxidases is necessary for iron efflux from stores via Ferroportin1 (Fpn1). Indeed, in absence of ceruloplasmin activity, iron accumulates in the liver causing anemia (Osaki et al., 1971; Harris et al., 1999; Kosman, 2010; Ganz, 2013). Therefore, the decreased oxidase activity we observed in Prnp0/0 mouse liver and serum is likely the cause of iron accumulation in the liver and decrease in the serum.


In Absence of the Cellular Prion Protein, Alterations in Copper Metabolism and Copper-Dependent Oxidase Activity Affect Iron Distribution
Comparison of Cu, Fe, oxidase activity, metal-binding protein expression levels in wild-type and PrPC- mouse liver at different ages. (A) The graph shows the ratio of Cu and Fe levels in Prnp0/0 and Prnp+/+ liver samples (P15, P90 N = 5; P30, P180, P365 N = 4). (B) The graph shows the levels of oxidase activity as U/mL in Prnp0/0 and Prnp+/+ liver (P15, P90, P180, P365 N = 4; P30 N = 5). (C) Representative Western blot images showing metal-binding protein levels in Prnp0/0 and Prnp+/+ liver samples. The constant level of the housekeeping protein (β-Actin) are also reported. (D) 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), N = 4. All error bars indicate SD; *p < 0.05; **p < 0.01.
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Figure 2: Comparison of Cu, Fe, oxidase activity, metal-binding protein expression levels in wild-type and PrPC- mouse liver at different ages. (A) The graph shows the ratio of Cu and Fe levels in Prnp0/0 and Prnp+/+ liver samples (P15, P90 N = 5; P30, P180, P365 N = 4). (B) The graph shows the levels of oxidase activity as U/mL in Prnp0/0 and Prnp+/+ liver (P15, P90, P180, P365 N = 4; P30 N = 5). (C) Representative Western blot images showing metal-binding protein levels in Prnp0/0 and Prnp+/+ liver samples. The constant level of the housekeeping protein (β-Actin) are also reported. (D) 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), N = 4. All error bars indicate SD; *p < 0.05; **p < 0.01.
Mentions: Incorporation of copper into Cp mainly occurs in the liver (Terada et al., 1995). Therefore, we measured copper content in Prnp0/0 and Prnp+/+ mouse liver during early development and adulthood, and expressed results as the ratio between Prnp0/0 and Prnp+/+ ion concentration values (Figure 2A). Results expressed in μg/mL are reported in Figures S1D,E. Similarly to serum, total copper content is not overall altered in Prnp0/0 mouse liver, though a decrease of about 30% occurs at the early stage P15 (Figure 2A). Our data show iron accumulation in the liver starting from P90 (Figure 2A), corresponding to the reduction of serum iron concentration (Figure 1A). As for serum Cp activity, liver oxidase activity is diminished in Prnp0/0 mice (Figure 2B). The activity of ferroxidases is necessary for iron efflux from stores via Ferroportin1 (Fpn1). Indeed, in absence of ceruloplasmin activity, iron accumulates in the liver causing anemia (Osaki et al., 1971; Harris et al., 1999; Kosman, 2010; Ganz, 2013). Therefore, the decreased oxidase activity we observed in Prnp0/0 mouse liver and serum is likely the cause of iron accumulation in the liver and decrease in the serum.

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.