FVB/NJ-Tg(ACTA1-PRKAG1*R70Q)3Lwit/LjgJ

Stock number: 027389
Product name: AMPK R70Qγ1 , γ1R70Q TG , γ1TG , AMPK R70Q gamma1 , muscle-specific constitutively active (CA) AMPK-γ1^R70Q , ACTA1-HA-R70Qγ1
Research areas: Cardiovascular Research, Cell Biology Research, Research Tools

Description

Muscle-specific, constitutively active (CA) AMPK-γ1^R70Q transgenic mice have the human skeletal muscle α-actin promoter directing expression of the human R70Qγ1 protein. This results in chronic AMPK activation predominantly in fully differentiated skeletal muscle with some expression in cardiac myocytes.

Detailed description

The ACTA1-HA-R70Qγ1 transgene has the human skeletal muscle α-actin promoter regions directing expression of a hemagglutinin epitope-tagged, human AMP-activated protein kinase non-catalytic γ1 variant made constitutively active by the R70Q amino acid substitution (AMPK-γ1^R70Q). Transgenic mice are also called HA-R70Qγ1, AMPK R70Qγ1, γ1R70Q TG, γ1TG or muscle-specific constitutively active (CA) AMPK-γ1^R70Q mice.
Hemizygous mice from founder line 3 have AMPK-γ1^R70Q expression predominantly in fully differentiated skeletal muscle, and some expression also observed in cardiac myocytes. This muscle-specific AMPK-γ1^R70Q expression results in chronic AMPK activation (~threefold higher than wildtype) and increased muscle glycogen content. Specifically, AMPK-γ1^R70Q mice exhibit a small, but significant, increase in muscle glycogen synthase (GSY) activity associated with an increase in the muscle expression of the liver isoform GSY2. The increase in glycogen content is accompanied by an increase in exercise capacity. Despite activated AMPK, basal p38 MAPK phosphorylation was not different than wildtype. Hemizygotes show significantly blunted muscle contraction-induced p38 MAPK phosphorylation compared to wildtype. In addition, the donating investigator reports that sedentary AMPK-γ1^R70Q mice show a 2.6-fold increase in type IIa/x fibers, a significant increase in hexokinase II and an ~25% increase in citrate synthase activity compared to wildtype, but had no further increases with exercise training.
Hemizygotes are viable and fertile, with normal locomotive activity, rate of oxygen consumption, rate of carbon dioxide generation, food consumption and water consumption.
Of note, when AMPK-γ1^R70Q mice are bred to also have the RyR1^Y524S mutation (Ryr1^tm1Slh), presence of the ACTA1-HA-R70Qγ1 transgene did not rescue the lethality of heat-sensitive RyR1^Y524S heterozygotes.

Development

The ACTA1-HA-R70Qγ1 transgene was designed by Dr. Lee A. Witters (Dartmouth Medical School) to have the human skeletal muscle α-actin promoter (~2.2 kbp of the 5prime region of ACTA1), a full-length human AMP-activated protein kinase non-catalytic γ1 cDNA sequence harboring the R70Q amino acid substitution (R70Qγ1) tagged at the 5prime end with a hemagglutinin (HA) epitope, all followed by the SV40 sequence. The 3.6 kbp transgene was microinjected into the pronuclei of FVB/NJ fertilized eggs. Founder animals were bred to wildtype FVB/NJ inbred mice to generate transgenic lines.

The donating investigator reports that, as described in the primary publication, hemizygous mice from founder line 3 were found to have approximately threefold increase in total γ1 gene copy number compared with wildtype mice. These mice were maintained on an FVB/NJ genetic background and then white hemizygous males were sent to Dr. Laurie J. Goodyear (Harvard Medical School) in 2002. There, hemizygotes were bred once to FVB/NJ inbred mice, and thereafter bred to wildtype (noncarrier) mice from the colony, wildtype (noncarrier) siblings and/or FVB/NJ inbred mice for several generations. In 2015, Dr. Goodyear sent white hemizygous males to The Jackson Laboratory Repository. Upon arrival, males were used to cryopreserve sperm. To establish the living mouse colony, an aliquot of the frozen sperm is used to fertilize oocytes from FVB/NJ inbred females (Stock No. 001800).

Allele

Symbol: Tg(ACTA1-PRKAG1*R70Q)3Lwit
Name: transgene insertion 3, Lee Witters
MGI accession ID: MGI:5637561
Generation method: Transgenic
Attributes: Inserted expressed sequence; Humanized sequence
Marker symbol: Tg(ACTA1-PRKAG1*R70Q)3Lwit
Marker name: transgene insertion 3, Lee Witters
Chromosome: UN
Strain of origin: FVB/NJ
Expressed genes: PRKAG1,protein kinase AMP-activated non-catalytic subunit gamma 1,human
Molecular note: The 3.6 kbp transgene contains a human skeletal muscle alpha-actin promoter (~2.2 kbp of the 5prime region of ACTA1), a full-length human AMP-activated protein kinase non-catalytic gamma1 (PRKAG1) cDNA sequence harboring the R70Q amino acid substitution tagged to the 5' end with a hemagglutinin (HA) epitope, all followed by the SV40 sequence. Immunoblot detects transgene expression in gastrocnemius, tibialis anterior, quadriceps and extensor digitorum longus muscles. There is little detectable expression in the soleus and low level expression in heart. Founder line 3 has approximately threefold increase in total gamma1 gene copy number compared with wildtype mice