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Premutation in the Fragile X Mental Retardation 1 (FMR1) Gene Affects Maternal Zn-milk and Perinatal Brain Bioenergetics and Scaffolding.

Napoli E, Ross-Inta C, Song G, Wong S, Hagerman R, Gane LW, Smilowitz JT, Tassone F, Giulivi C - Front Neurosci (2016)

Bottom Line: Given that the most significant effects were observed at the end of the lactation period, we hypothesized that KI milk might have a role at compounding the deleterious effects on the FMR1 genetic background.A highly significant milk type × genotype interaction was observed for all three-brain regions, being cortex the most influenced.Finally, lower milk-Zn levels were recorded in milk from lactating women carrying the premutation as well as other Zn-related outcomes (Zn-dependent alkaline phosphatase activity and lactose biosynthesis-whose limiting step is the Zn-dependent β-1,4-galactosyltransferase).

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biosciences, School of Veterinary Medicine Davis, CA, USA.

ABSTRACT
Fragile X premutation alleles have 55-200 CGG repeats in the 5' UTR of the FMR1 gene. Altered zinc (Zn) homeostasis has been reported in fibroblasts from >60 years old premutation carriers, in which Zn supplementation significantly restored Zn-dependent mitochondrial protein import/processing and function. Given that mitochondria play a critical role in synaptic transmission, brain function, and cognition, we tested FMRP protein expression, brain bioenergetics, and expression of the Zn-dependent synaptic scaffolding protein SH3 and multiple ankyrin repeat domains 3 (Shank3) in a knock-in (KI) premutation mouse model with 180 CGG repeats. Mitochondrial outcomes correlated with FMRP protein expression (but not FMR1 gene expression) in KI mice and human fibroblasts from carriers of the pre- and full-mutation. Significant deficits in brain bioenergetics, Zn levels, and Shank3 protein expression were observed in the Zn-rich regions KI hippocampus and cerebellum at PND21, with some of these effects lasting into adulthood (PND210). A strong genotype × age interaction was observed for most of the outcomes tested in hippocampus and cerebellum, whereas in cortex, age played a major role. Given that the most significant effects were observed at the end of the lactation period, we hypothesized that KI milk might have a role at compounding the deleterious effects on the FMR1 genetic background. A higher gene expression of ZnT4 and ZnT6, Zn transporters abundant in brain and lactating mammary glands, was observed in the latter tissue of KI dams. A cross-fostering experiment allowed improving cortex bioenergetics in KI pups nursing on WT milk. Conversely, WT pups nursing on KI milk showed deficits in hippocampus and cerebellum bioenergetics. A highly significant milk type × genotype interaction was observed for all three-brain regions, being cortex the most influenced. Finally, lower milk-Zn levels were recorded in milk from lactating women carrying the premutation as well as other Zn-related outcomes (Zn-dependent alkaline phosphatase activity and lactose biosynthesis-whose limiting step is the Zn-dependent β-1,4-galactosyltransferase). In premutation carriers, altered Zn homeostasis, brain bioenergetics and Shank3 levels could be compounded by Zn-deficient milk, increasing the risk of developing emotional and neurological/cognitive problems and/or FXTAS later in life.

No MeSH data available.


Related in: MedlinePlus

Correlation between mitochondrial outcomes and FMRP or FMR1 expression levels. The correlation between FMRP and FMR1 expression with a mitochondrial outcome (i.e., coupling between ATP synthesis and electron transfer or RCRu) was carried out using human primary dermal from controls, premutation carriers (105–180 CGG), and full mutation carriers (>200 CGG repeats). RCRu, FMR1, and FMRP levels were expressed as percentage of control values. Data are shown as mean ± SEM for controls, permutation, and full mutation carriers.
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Figure 4: Correlation between mitochondrial outcomes and FMRP or FMR1 expression levels. The correlation between FMRP and FMR1 expression with a mitochondrial outcome (i.e., coupling between ATP synthesis and electron transfer or RCRu) was carried out using human primary dermal from controls, premutation carriers (105–180 CGG), and full mutation carriers (>200 CGG repeats). RCRu, FMR1, and FMRP levels were expressed as percentage of control values. Data are shown as mean ± SEM for controls, permutation, and full mutation carriers.

Mentions: As a proof of concept, to test whether FMR1 gene or FMRP protein expression influence mitochondrial outcomes, we turned to human primary skin fibroblasts from premutation and full mutation carriers, as commercially available FMR1 KO mice are on a different genetic background (FVB129 or FVB129/NJ) than the ones used in this study (C57BL/6J) making the comparison invalid [see as an example for Huntington's disease, another triplet-nucleotide repeat disease (Menalled et al., 2009)]. In addition, KO mice may not recapitulate the full mutation seen in humans, since humans show no expression of FMRP over time as a result of a repeat-mediated gene silencing sometime during early development (Pieretti et al., 1991; Sutcliffe et al., 1992). To this end, FMR1 and FMRP expression were assessed in fibroblasts from controls, premutation (105–180 CGG matching the range of CGG in KI mice), and full mutation (>200 CGG repeats) carriers (Figure 4). Fibroblasts from premutation carriers showed normal or marginally elevated FMR1 transcript levels with ~50% FMRP levels of controls; in contrast, fibroblasts from full mutation carriers showed no gene or protein expression. When the link between FMR1 and FMRP expression and bioenergetics (as judged by coupling between ATP synthesis and electron transfer, RCRu) was investigated in these samples, a strong direct correlation was observed between RCRu and FMRP levels but not FMR1 mRNA levels (Figure 4).


Premutation in the Fragile X Mental Retardation 1 (FMR1) Gene Affects Maternal Zn-milk and Perinatal Brain Bioenergetics and Scaffolding.

Napoli E, Ross-Inta C, Song G, Wong S, Hagerman R, Gane LW, Smilowitz JT, Tassone F, Giulivi C - Front Neurosci (2016)

Correlation between mitochondrial outcomes and FMRP or FMR1 expression levels. The correlation between FMRP and FMR1 expression with a mitochondrial outcome (i.e., coupling between ATP synthesis and electron transfer or RCRu) was carried out using human primary dermal from controls, premutation carriers (105–180 CGG), and full mutation carriers (>200 CGG repeats). RCRu, FMR1, and FMRP levels were expressed as percentage of control values. Data are shown as mean ± SEM for controls, permutation, and full mutation carriers.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4835505&req=5

Figure 4: Correlation between mitochondrial outcomes and FMRP or FMR1 expression levels. The correlation between FMRP and FMR1 expression with a mitochondrial outcome (i.e., coupling between ATP synthesis and electron transfer or RCRu) was carried out using human primary dermal from controls, premutation carriers (105–180 CGG), and full mutation carriers (>200 CGG repeats). RCRu, FMR1, and FMRP levels were expressed as percentage of control values. Data are shown as mean ± SEM for controls, permutation, and full mutation carriers.
Mentions: As a proof of concept, to test whether FMR1 gene or FMRP protein expression influence mitochondrial outcomes, we turned to human primary skin fibroblasts from premutation and full mutation carriers, as commercially available FMR1 KO mice are on a different genetic background (FVB129 or FVB129/NJ) than the ones used in this study (C57BL/6J) making the comparison invalid [see as an example for Huntington's disease, another triplet-nucleotide repeat disease (Menalled et al., 2009)]. In addition, KO mice may not recapitulate the full mutation seen in humans, since humans show no expression of FMRP over time as a result of a repeat-mediated gene silencing sometime during early development (Pieretti et al., 1991; Sutcliffe et al., 1992). To this end, FMR1 and FMRP expression were assessed in fibroblasts from controls, premutation (105–180 CGG matching the range of CGG in KI mice), and full mutation (>200 CGG repeats) carriers (Figure 4). Fibroblasts from premutation carriers showed normal or marginally elevated FMR1 transcript levels with ~50% FMRP levels of controls; in contrast, fibroblasts from full mutation carriers showed no gene or protein expression. When the link between FMR1 and FMRP expression and bioenergetics (as judged by coupling between ATP synthesis and electron transfer, RCRu) was investigated in these samples, a strong direct correlation was observed between RCRu and FMRP levels but not FMR1 mRNA levels (Figure 4).

Bottom Line: Given that the most significant effects were observed at the end of the lactation period, we hypothesized that KI milk might have a role at compounding the deleterious effects on the FMR1 genetic background.A highly significant milk type × genotype interaction was observed for all three-brain regions, being cortex the most influenced.Finally, lower milk-Zn levels were recorded in milk from lactating women carrying the premutation as well as other Zn-related outcomes (Zn-dependent alkaline phosphatase activity and lactose biosynthesis-whose limiting step is the Zn-dependent β-1,4-galactosyltransferase).

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biosciences, School of Veterinary Medicine Davis, CA, USA.

ABSTRACT
Fragile X premutation alleles have 55-200 CGG repeats in the 5' UTR of the FMR1 gene. Altered zinc (Zn) homeostasis has been reported in fibroblasts from >60 years old premutation carriers, in which Zn supplementation significantly restored Zn-dependent mitochondrial protein import/processing and function. Given that mitochondria play a critical role in synaptic transmission, brain function, and cognition, we tested FMRP protein expression, brain bioenergetics, and expression of the Zn-dependent synaptic scaffolding protein SH3 and multiple ankyrin repeat domains 3 (Shank3) in a knock-in (KI) premutation mouse model with 180 CGG repeats. Mitochondrial outcomes correlated with FMRP protein expression (but not FMR1 gene expression) in KI mice and human fibroblasts from carriers of the pre- and full-mutation. Significant deficits in brain bioenergetics, Zn levels, and Shank3 protein expression were observed in the Zn-rich regions KI hippocampus and cerebellum at PND21, with some of these effects lasting into adulthood (PND210). A strong genotype × age interaction was observed for most of the outcomes tested in hippocampus and cerebellum, whereas in cortex, age played a major role. Given that the most significant effects were observed at the end of the lactation period, we hypothesized that KI milk might have a role at compounding the deleterious effects on the FMR1 genetic background. A higher gene expression of ZnT4 and ZnT6, Zn transporters abundant in brain and lactating mammary glands, was observed in the latter tissue of KI dams. A cross-fostering experiment allowed improving cortex bioenergetics in KI pups nursing on WT milk. Conversely, WT pups nursing on KI milk showed deficits in hippocampus and cerebellum bioenergetics. A highly significant milk type × genotype interaction was observed for all three-brain regions, being cortex the most influenced. Finally, lower milk-Zn levels were recorded in milk from lactating women carrying the premutation as well as other Zn-related outcomes (Zn-dependent alkaline phosphatase activity and lactose biosynthesis-whose limiting step is the Zn-dependent β-1,4-galactosyltransferase). In premutation carriers, altered Zn homeostasis, brain bioenergetics and Shank3 levels could be compounded by Zn-deficient milk, increasing the risk of developing emotional and neurological/cognitive problems and/or FXTAS later in life.

No MeSH data available.


Related in: MedlinePlus