New Mouse Models Developed of Rare Mutation Seen in Some Patients

New models fill 'important gap' for future study

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by Steve Bryson, PhD |

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Researchers have developed and characterized mouse models of Friedreich’s ataxia (FA) carrying a rare disease-causing mutation, a study reports.

The rare FXN gene mutation, dubbed G130V, is found in some FA patients alongside the common gene expansion defects seen in most cases. These patients experience less severe symptoms and slower disease progression, despite producing very low levels of frataxin protein.

Although researchers found that mice with two equivalent mutations had undetectable frataxin, grew smaller, and had low physical endurance and activity, their motor coordination remained intact.

According to researchers, this FA mouse model fills an important gap between milder expansion models and those with severe frataxin deficiency and can be used in therapeutic approaches, especially those relying on frataxin replacement.

The study, “Neurobehavioral deficits of mice expressing a low level of G127V mutant frataxin,” was published in the journal Neurobiology of Disease.

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Most cases of FA are caused by defects in both copies of the FXN gene, each inherited from one parent. Such defects lead to the production of two extended genes containing extra GAA repeats in their DNA sequence (G stands for guanine and A for adenosine, two DNA building blocks).

As a result, people with FA have low levels of the FXN-encoded frataxin protein, which disrupts energy generation within cells, particularly those of the muscles and nervous system.

In rare cases, some FA patients have an expanded GAA repeat sequence in one FXN gene copy and a G130V mutation in the other copy.

US researchers engineer multiple mouse models with various genotypes

To further understand the impact of this rare mutation, researchers in the U.S. engineered and examined multiple FA mouse models with various genotypes — or combinations of GAA expansion and G127V mutations (equivalent to the human G130V).

One FA mouse model carried two G127V mutations, one had a GAA expansion and a G127V mutation (GAA/G127V), and another had one GAA expansion and a deleted Fxn gene (GAA/Fxn deletion).

As early as 3 months of age, mice with two G127V mutations were significantly smaller than the other models, and also developed mild curvature of the spine, which progressed to severe at one year, similar to the GAA/Fxn deletion model.

Hindlimb clasping is a hallmark feature in ataxia mouse models, where animals involuntarily retract one or both hindlimbs toward their abdomen when held by the tail.

While the double G127V mice did not show this behavior, some mice of the GAA/G127V model developed mild but persistent clasping at six months. By one year, however, all models showed some clasping behavior, “suggesting that age rather than genotype influenced the presentation,” the team wrote.

All models showed similar levels of Fxn gene expression (activity) compared with unaffected mice, except for GAA/Fxn deletion mice, which showed a 40%–50% reduction. By contrast, at three months and one year, frataxin protein levels were significantly reduced in brain, spinal, and heart tissue of double G127V, GAA/G127V, and GAA/Fxn deletion mice.

Notably, while frataxin levels were reduced by up to 40% in GAA/G127V and GAA/Fxn deletion mice, little to no frataxin was detected in double G127V mutant samples. Frataxin G127V protein was also found to localize to the mitochondria but at 1.7% relative to healthy samples.

Mice tested for locomotor activity, strength, and gait

Mice were then subjected to various tests for locomotor activity (open field), strength (forelimb grip strength), and gait.

At three months, double G127V male mice traveled shorter distances more slowly in the open field test than controls. This was sustained versus other models, but male double G127V mice moved faster after one year. Female double mutants, by comparison, also showed reduced speed than other models at three months, which was sustained at later timepoints.

Forelimb grip strength steadily declined with age for both sexes across all models, with significantly reduced strength noted in double G127V mutant male and female mice.

Stride length, the distance between the successive placement of the same paw, was significantly shorter for double G127V mutant and GAA/G127V male mice in the forelimbs and hindlimbs. Female double G127V mutant mice also had small stride lengths in all limbs, while GAA/G127V animals had significantly longer front limb stride lengths than controls at 3 months of age. Paw coordination was generally unaffected by either genotype.

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Genes altered in double G127V mutant mice were related to ribosomes (protein production), proteasomes (protein recycling), and oxidative phosphorylation (energy production). By comparison, altered GAA/G127V genes were involved in signaling, addiction, and synapses, the point where nerve cells communicate.

Gene expression patterns in the oxidative phosphorylation and ribosome pathways demonstrated that nearly all genes altered in double mutant mice were significantly less active, “indicating a severe metabolic imbalance caused by the Fxn G127V missense mutation at the cellular level,” the team noted.

“The [double G127V] mouse model fills an important gap in [Friedreich’s ataxia] animal models between … mild GAA repeat expansion models and severely impaired mice caused by rapid and drastic depletion of Fxn,” the researchers concluded. “It represents a model of severe and inherited frataxin decrease that displays moderate and traceable phenotypes.”

“These characteristics make the [double G127V] mouse a good model to study consequences of frataxin deficiency and a tool for development of therapeutic approaches, especially those relying on frataxin replacement or supplementation,” the researchers added.

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