TG-hATXN3(85Q) Mice

Catalog Number: C001397

Strain Name: C57BL/6JCya-Tg(hATXN3*85Q)/Cya

Genetic Background: C57BL/6JCya

 

Strain Description

The ATXN3 gene encodes Ataxin 3, a protein primarily responsible for intracellular protein degradation and involved in various cellular processes including DNA repair and autophagy. The ATXN3 gene is widely expressed in the human brain and other tissues, with particularly high expression levels in the cerebellum and spinal cord [1]. Spinocerebellar Ataxia type 3 (SCA3), also known as Machado-Joseph Disease (MJD), is a progressive neurodegenerative disorder characterized clinically by motor coordination impairment (cerebellar ataxia), bulbar, pyramidal, and extrapyramidal dysfunction, and may be accompanied by peripheral neuropathy or ophthalmoplegia [2]. SCA3 is the most common dominantly inherited ataxia, caused by an abnormal expansion of CAG repeat sequences in the ATXN3 gene. This expansion leads to the formation of an elongated polyglutamine (polyQ) domain in the Ataxin 3 protein, subsequently causing protein aggregation and dysfunction of the ubiquitin-proteasome system. Among polyglutamine diseases, the prevalence of SCA3 is second only to Huntington's disease (HD). The number of CAG repeats in the ATXN3 gene of healthy individuals typically ranges from 12 to 44, whereas affected individuals with SCA3 have 56 to 87 repeats. Individuals with 45 to 55 repeats may exhibit incomplete penetrance of SCA3 symptoms. Similar to other polyglutamine diseases, the length of the CAG repeat is negatively correlated with the age of onset and positively correlated with the severity of SCA3 [2-3]

This strain represents a SCA3 disease model generated through transgenic technology, expressing a human ATXN3 gene carrying approximately 85 CAG repeats (Q). The number of CAG repeats (Q) in the human ATXN3 gene in this model is associated with the more severe forms of SCA3. Preliminary research data indicate that 2-month-old TG-hATXN3(85Q) mice exhibit a significant increase in stride width and a shortened latency to fall in the rotarod test, suggesting impairments in motor coordination and activity. Therefore, this strain can be utilized for research on the ubiquitin-proteasome system and the pathogenic mechanisms of SCA3, as well as for the screening, development, and evaluation of targeted therapeutic drugs.

 

Strain Strategy

The mouse model was generated through transgenic (TG) technology by integrating a human ATXN3 gene carrying approximately 85 CAG repeats (Q) into the mouse genome.

 

Applications

  • Research on the ubiquitin-proteasome system and the pathological mechanisms of SCA3.
  • Screening, development, and evaluation of therapeutic drugs for SCA3.

 

Validation Data

1. Protein Expression

a. Comparison of expression levels of related strains

Figure 1. Western Blot results of the cerebellum and spinal cord of 8-week-old male B6-hATXN3 (Product No.: C001398), TG-hATXN3(85Q) (Product No.: C001397), B6-TG(ATXN3 - 84Q) (Product No.: C001434) and wild-type (WT) mice.

①According to the detection method provided in the instruction manual of the human-specific antibody reagent, the theoretical molecular weight of ATXN3 is predicted to be 42KD. Due to the high homology of protein sequences between wild-type mice and humans, corresponding bands were detected in WT and humanized mice. Compared with WT, the molecular weight of the target band detected in B6-hATXN3 mice is larger because the amino acid sequence of human ATXN3 (361aa) is longer than that of mouse Atxn3 (355aa).

②Bands appeared around 65-70KD in transgenic mice TG-hATXN3(85Q) and B6-TG(ATXN3-84Q). This is because different numbers of CAG repeat (Q) sequences were introduced into these two types of mice, increasing the molecular weight of the target band. In addition, both transgenic mice are heterozygotes, and bands of human origin (about 65-70KD) and murine origin (about 42KD) can be detected simultaneously. Therefore, the WB results show a double-band characteristic. The band of TG-hATXN3(85Q) mice is slightly higher than that of B6-TG(ATXN3-84Q) mice, but its expression level is slightly lower.

 

b. Comparison of gender differences

Figure 2. Western Blot results of the cerebellum and spinal cord of 8-week-old TG-hATXN3(85Q) mice and wild-type (WT) mice.

①Due to the high homology of protein sequences between wild-type mice and humans, corresponding bands were detected in both WT and TG-hATXN3(85Q) mice.

②A band appeared around 70KD in TG-hATXN3(85Q) mice. This is because the human ATXN3 sequence introduced into this strain carries a specific number of CAG repeat (Q) sequences, which leads to an increase in the molecular weight of the target band. In addition, TG-hATXN3(85Q) mice are heterozygotes, allowing the simultaneous detection of bands of human and murine origins, thus exhibiting the characteristic of double bands.

③There was no significant difference in the expression level of ATXN3 protein between female and male TG-hATXN3(85Q) mice.

 

2. Gait Test: Stride Width

Figure 3. The forelimb/hindlimb stride width of 2-month-old (2M) mice in the gait test was measured. Statistical comparisons were conducted using unpaired t-test, with p < 0.05 indicating significance. 

Indications: For male and sex-mixed comparisons, TG-hATXN3(85Q) exhibited a significant rise in forelimb/hindlimb stride width, implying impairments in locomotor coordination.

Mouse numbers: TG-hATXN3(85Q) mice, 10 females and 10 males, total 20; Wild-type mice, 12 females and 12 males, total 24; the same applies below.

 

3. Gait Test: Left Forelimb/Hindlimb Stride Length

Figure 4. The left forelimb/hindlimb stride length of 2M mice in the gait test was measured. Statistical comparisons were conducted using unpaired t-test, with p < 0.05 indicating significance. 

Indications: No significant alterations in left forelimb/hindlimb stride length across all models.

 

4. Gait Test: Right Forelimb/Hindlimb Stride Length

Figure 5. The right forelimb/hindlimb stride length of 2M mice in the gait test was measured. Statistical comparisons were conducted using unpaired t-test, with p < 0.05 indicating significance. 

Indications: No significant alterations in right forelimb/hindlimb stride length across all models.

 

5. Rotarod Test: Latency to Fall

Figure 6. The latency to fall of 2M mice in the rotarod was measured. Statistical comparisons were conducted using Mann-Whitney test, with p < 0.05 indicating significance.

Indications: For female and sex-mixed comparisons, TG-hATXN3(85Q) exhibited a significant decrease in latency to fall on rotarod test, indicating impairments in locomotor activity and coordination.

 

References
[1]Takiyama Y, Nishizawa M, Tanaka H, Kawashima S, Sakamoto H, Karube Y, Shimazaki H, Soutome M, Endo K, Ohta S, et al. The gene for Machado-Joseph disease maps to human chromosome 14q. Nat Genet. 1993 Jul;4(3):300-4.
[2]Paulson H. Machado-Joseph disease/spinocerebellar ataxia type 3. Handb Clin Neurol. 2012;103:437-49.
[3]McLoughlin HS, Moore LR, Paulson HL. Pathogenesis of SCA3 and implications for other polyglutamine diseases. Neurobiol Dis. 2020 Feb;134:104635.