This Fmr1 CGG repeat expansion premutation knock-in (Fmr1 CGG KI) mouse model is useful for studying Fragile X Tremor/Ataxia Syndrome (FXTAS).
David L Nelson, Baylor College of Medicine
Rob Willemsen, Erasmus Medical Center
The human fragile X mental retardation 1 (FMR1) gene on the X chromosome contains an unstable CGG repeat in its 5' untranslated region. While repeat length in the normal population is polymorphic (5–54 CGG repeats), CGG repeat expansions greater than 200 (i.e., the full mutation) have hypermethylation that results in FMR1 gene silencing and fragile X syndrome. Individuals with CGG repeat expansions of 55–200 (premutation CGG or preCGG) can develop the progressive neurodegenerative disease fragile X-associated tremor/ataxia syndrome (FXTAS).
The Fmr1 CGG repeat expansion premutation knock-in allele (Fmr1 CGG KI) was generated by substituting the endogenous mouse (CGG)8 with a human (CGG)98. Similar to humans, germline passage of this allele in mice can result in contraction and/or expansion of the meiotically unstable CGG repeat element; resulting in a broad range of allele sizes (<70 to >300 CGG repeats).
The Jackson Laboratory will attempt to have Fmr1 CGG KI mice with >90 CGG repeats [(CGG)90+] available.
In general, Fmr1 CGG KI mice show elevated Fmr1 mRNA levels, defects in neuronal morphology and migration, and ubiquitin-positive neuronal inclusions throughout the brain. Fmr1 CGG KI mice exhibit aberrant behavior, including mild learning deficits and increased anxiety. The donating investigator reports that homozygous females and hemizygous males are viable and fertile, and when bred together they observe no breeding abnormalities (litter size/frequency, nursing, etc.). The phenotype of mice with specific repeat lengths are described below.
Hemizygous males with (CGG)102-110 exhibit ubiquitin positive intranuclear neuronal inclusions.
Hemizygous males with (CGG)106-123 exhibit abnormal learning/memory/conditioning (52 weeks old), impaired coordination (52 weeks old and 72 weeks old), decreased grip strength (52 weeks old) and abnormal behavior (72 weeks old). However, gait, activity levels and exploration levels are normal at 72 weeks of age.
Homozygous females and hemizygous males with up to (CGG)200 are viable and fertile.
In Fmr1 CGG KI mice, the CGG repeat number is subject to germline and somatic instability, and may expand or contract. When using mouse models with unstable CGG repeat length, it is strongly recommended the CGG repeat number be quantified in all the experimental animals; all animals in all experimental groups should carry comparable CGG repeat sizes.
The Fmr1 CGG repeat expansion premutation knock-in allele (Fmr1 CGG KI) was created by Dr. Ben A. Oostra (Erasmus University). The targeting vector was designed to replace the endogenous 8 CGG repeat sequence [(CGG)8] in the promoter of the fragile X mental retardation syndrome 1 gene (Fmr1) on the X chromosome with a human 98 CGG repeat sequence [(CGG)98] and also place a loxP-flanked neo cassette into intron 1. The construct was electroporated into 129P2/OlaHsd-derived E14 embryonic stem (ES) cells. Correctly targeted ES cells were injected into recipient blastocysts, and chimeras were bred with FVB and/or C57BL/6 to establish the colony.
At some point, the (CGG)98/neo animals were bred with germline Cre recombinase-expressing mice (undisclosed genetic background background) to delete the neo cassette. The resulting mice are called (CGG)98/neo- (or Fmr1 CGG KI).
In ~2007, Fmr1 CGG KI mice on a C57BL/6J;FVB/N genetic background with black coat color were sent to Dr. David L. Nelson (Baylor College of Medicine). There, Dr. Nelson backcrossed the colony one generation to C57BL/6J females.
Fmr1 CGG KI males with a donor-reported ~130 CGG repeats [(CGG)~130] and black coat color were sent to The Jackson Laboratory Repository in 2016. Upon arrival, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6J oocytes (Stock No. 000664).
|Allele Name||targeted mutation 2, Ben Oostra|
|Allele Type||Targeted (Humanized sequence)|
|Gene Symbol and Name||Fmr1, fragile X mental retardation 1|
|Strain of Origin||129P2/OlaHsd|
|Molecular Note||The mouse promoter 8 CGG repeat sequence was replaced with a human 98 CGG repeat and a floxed neo cassette via homologous recombination. The targeted allele was verified by PCR analysis. Germline passage of this allele results in contraction and expansion of the CGG repeats.|
The Fmr1 locus is on the X chromosome. When maintaining a live colony, wildtype females from the colony may be bred with hemizygous males. Alternatively, heterozygous females may be bred with wildtype males from the colony. The donating investigator reports that homozygous females and hemizygous males are viable and fertile, and when bred together they observe no breeding abnormalities (litter size/frequency, nursing, etc.).
When using the Fmr1 CGG KI mouse strain in a publication, please cite the originating article(s) and include JAX stock #024102 in your Materials and Methods section.