C57BL/6N-Mdga2^{Tg(Prnp-PFN1*G118V)838Kiaei}/J

Stock number: 030568
Product name: PrP-hPFN1^G118V
Research areas: Cell Biology Research, Neurobiology Research, Research Tools

Description

PrP-PFN1^G118V transgenic mice have expression of an untagged human profilin 1 cDNA sequence encoding the G118V mutant protein (hPFN1^G118V) associated with amyotrophic lateral sclerosis (ALS; Lou Gehrig's Disease) and directed to brain and spinal cord by the mouse prion protein promoter. PrP-PFN1^G118V mice can exhibit a long symptomatic period (~120-200 days of age on average) of ALS characteristics. Along with the similarly designed human PFN1 wildtype-expressing transgenic mice (PrP-PFN1^WT; Stock No. 030570), these mice may be useful in studying cytoskeletal dysfunction as a neurodegenerative factor in ALS, as well as ALS neurotoxicity, proteinopathy and axonal degeneration.

As of October 2018, sequencing analysis identified the Prp-hPFN1^G118V transgene inserted on chromosome 12 causing a 151 kb deletion in Mdga2 (exons 2-3).

Detailed description

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder characterized by the loss of upper and lower motor neurons. Affected individuals develop progressive muscle weakness and atrophy, eventually leading to death due to respiratory failure. Identification of mutations in the profilin1 gene (PFN1) in human familial ALS patients focused attention on cytoskeletal dysfunction as a neurodegenerative factor in ALS.

Prp-hPFN1^G118V transgenic mice express an untagged human profilin 1 cDNA sequence modified to have the ALS-linked G118V mutation (hPFN1^G118V) under the direction of the mouse prion protein promoter (Prnp; PrP). As expected for the PrP promoter sequences, transgene expression is confined primarily to central nervous system (brain and spinal cord), with lower expression in skeletal muscle. No expression is reported in the liver, and expression levels in other tissues have not been examined to date (July 2017).

As of October 2018, sequencing analysis identified the Prp-hPFN1^G118V transgene inserted on chromosome 12 causing a 151 kb deletion in Mdga2 (exons 2-3). While this may be expected to result in a Mgda2 knock-out allele, the MGDA2 levels have not been characterized to date. Mice hemizygous for the Prp-hPFN1^G118V transgene are expected to have retained the Mgda2 wildtype allele on the homologous chromosome 12. The Prp-hPFN1^G118V transgene copy number is not yet known (October 2018).

The phenotype summarized below is for "highly-expressing" Prp-hPFN1^G118V transgenic mice on a C57BL/6N genetic background and maintained after 30 days of age with a diet restriction of 4-5 grams of standard chow per day per mouse (all as described in Fil et al. 2017 Hum Mol Genet 26:686). Original characterization of Prp-hPFN1^G118V transgenic mice identified total PFN1 protein levels in spinal cord at ~5-fold greater levels compared to endogenous mouse PFN1 protein levels in non-transgenic mice. The transgene insertion site(s) and copy number were not determined at that time (see strain Development section for further details, if any). The total PFN1 protein levels in the spinal cords were similar at all time-points tested (from 50 to 209 days of age). The transgene copy number and/or expression levels have not been reevaluated to date (July 2017).
Prp-hPFN1^G118V transgenic mice with these expression levels exhibit many key clinical and pathological characteristics observed in human ALS, including loss of lower (ventral horn) and upper motor neurons (corticospinal motor neurons in layer V), mutant profilin 1 aggregation, abnormally ubiquitinated proteins, reduced choline acetyltransferase (ChAT) enzyme expression, fragmented mitochondria (resembling Wallerian-like degeneration), glial cell activation, muscle atrophy, weight loss and reduced survival.
Specifically, hemizygous Prp-hPFN1^G118V mice develop progressively deteriorating motor dysfunction (average onset 120-130 days of age) and rapid progression to end-stage disease (average 165-210 days of age). There is no obvious difference in the time of disease onset between hemizygous females and hemizygous males, although the donating investigator reports that hemizygous females generally have shorter life span than hemizygous males. This long symptomatic period may enable ALS researchers to utilize this model to address the neurotoxicity, proteinopathy, cytoskeletal defects and axonal degeneration caused by G118V mutant PFN1.
The donating investigator reports Prp-hPFN1^G118V hemizygous mice are viable and fertile, with the normal breeding characteristics and normal littler sizes between 60-130 days of age. Additionally, they report hemizygous males breed well, but hemizygous females can be poor breeders even prior to exhibiting any neuromuscular phenotype (average onset 120-130 days). The donating investigator has not attempted to generate homozygous mice to date (July 2017), but expects that, compared to hemizygotes, the homozygotes would have higher hPFN1^G118V expression levels and an exacerbation of phenotype.

Importantly, when maintained ad libitum on standard chow, the C57BL/6N-congenic Prp-hPFN1^G118V mice can become obese and fail to exhibit characteristics of ALS. To prevent the tendency to become overweight, it is recommended to maintain these mice after 30 days of age with a diet restriction of 4-5 grams of standard chow per day per mouse. [Fil et al. 2017 Hum Mol Genet 26:686]

Development

The PrP-hPFN1^G118V transgene was designed by Dr. Mahmoud Kiaei (University of Arkansas for Medical Sciences). The PrP-PFN1^G118V transgene has an untagged human profilin 1 (PFN1) cDNA sequence (420 bp) encoding the G118V mutant protein (hPFN1^G118V; associated with familial amyotrophic lateral sclerosis [fALS]) inserted between exon 2 and exon 3 of mouse prion protein (PrP or Prnp) gene at an unique XhoI site in the MoPrP.XhoI plasmid vector (ATCC#JHU-2). The resulting ~14 kbp PrP-PFN1^G118V construct was microinjected into pronuclei of C57BL/6NCrl embryos (C57BL/6NCrl eggs fertilized by C57BL/6NCrl sperm). Transgenic founder mice were bred to C57BL/6NCrl mice. Founder line 838 was identified with total PFN1 protein levels in spinal cord at ~5-fold greater levels compared to endogenous mouse PFN1 protein levels in non-transgenic mice. The PrP-PFN1^G118V line 838 transgenic colony was maintained by breeding wild-type (noncarrier) females from the colony with hemizygous males for several generations. In 2017, transgenic males with black coat color were sent to The Jackson Laboratory Repository. Upon arrival, sperm was cryopreserved. To generate our live colony, an aliquot of the frozen sperm was used to fertilize oocytes from C57BL/6NJ mice (Stock No. 005304). Thereafter, the colony was maintained by breeding hemizygous mice with wild-type (noncarrier) mice from the colony or to C57BL/6NJ mice (Stock No. 005304).
As of October 2018, sequencing analysis identified the Prp-hPFN1^G118V transgene inserted on chromosome 12 causing a 151 kb deletion in Mdga2 (exons 2-3). While this may be expected to result in a Mgda2 knock-out allele, the MGDA2 levels have not been characterized to date. Mice hemizygous for the Prp-hPFN1^G118V transgene are expected to have retained the Mgda2 wildtype allele on the homologous chromosome 12. The Prp-hPFN1^G118V transgene copy number is not yet known (October 2018).