STOCK Dctn1^tm2.1Cai/J

Stock number: 032428
Product name: Dctn1^LoxP/
Research areas: Neurobiology Research, Research Tools

Description

Dctn1^LoxP/ mice have loxP sites flanking exons 2-4 of the dynactin 1 gene. Cre recombinase-mediated removal of the floxed sequence results in continued expression of the p135 subunit, but complete loss of p150^Glued subunit expression. These mice may be useful in studying the dynactin macromolecular complex in motor neuron disease and other neurodegenerative disorders.

Detailed description

Dynactin p150^Glued, the largest subunit of the dynactin macromolecular complex, binds to both microtubules and tubulin dimers through the N-terminal cytoskeleton-associated protein and glycine-rich (CAP-Gly) and basic domains, and serves as an anti-catastrophe factor in stabilizing microtubules in neurons. In addition to p150^Glued, DCTN1 also encodes p135 and other short splicing variants. The p135 lacks the coding exons 2 to 5, resulting in a complete loss of the CAP-Gly domain and a large portion of basic domain. The p150^Glued is expressed in all types of mammalian cells, while p135 is more abundant in neurons. Dynein/dynactin is required for the mitosis as germline deletion of p150^Glued caused early embryonic lethality and apoptosis in p150^Glued knock-out mice. However, p135 can compensate for the most dynactin activity in p150^Glued-deficient post-mitotic cells.

These Dctn1^LoxP/ mice have loxP sites flanking Dctn1 exons 2-4 (encoding the complete CAP-Gly domain and partial basic domain). Mice heterozygous or homozygous for this floxed allele (Dctn1^LoxP/+ or Dctn1^LoxP/LoxP, respectively) are viable and fertile, with normal fertility and no gross physical or behavioral abnormalities.

Upon Cre recombinase exposure, the deletion of Dctn1 exons 2-4 results in continued expression of the p135 subunit, but complete loss of p150^Glued subunit expression.

For example, when Dctn1^LoxP/ are bred to Thy1-Cre transgenic mice (Stock No. 006143), the resulting bi-genic conditional knock-out mice (p150^Glued cKO, also called Dctn1^LoxP/LoxP; Thy1-Cre) allow for the downstream motor behavioral and neuropathological studies. Specifically, the p150^Glued cKO mice did not show any obvious developmental phenotypes, but exhibited impairments in motor coordination and rearing after 12 months of age. Around 20% loss of spinal motor neurons was found in the lumbar spinal cord of 18-month-old p150^Glued cKO mice, in company with increased gliosis, neuromuscular junction (NMJ) disintegration and muscle atrophy. By contrast, no obvious degeneration of corticospinal motor neurons, striatal neurons, midbrain dopaminergic neurons, cerebellar granule cells or Purkinje cells was observed for p150^Glued cKO mice. [Yu et al. 2018 Mol Neurodegener. 13:10 (PMID:29490687)]

Furthermore, when Dctn1^LoxP/ are bred to CAG-Cre/Esr1 transgenic mice (Stock No. 004682) and exposed to tamoxifen, the donating investigator reports that the resulting mice homozygous for the global exons 2-4 knock-out allele (Dctn1^Δ2-4/Δ2-4) exhibit early embryonic lethality with 100% penetrance - a similar phenotype as their other mouse line with the global exons 2-3 knock-out allele (Dctn1^Δ2-3 ; see Lai et al. 2007 J Neurosci. 27:13982 (PMID:18094236)).

 

Development

The Dctn1^LoxP/ (floxed exons 2-4) allele was created by Dr. Huaibin Cai (NIA/NIH). First, a targeting vector was designed to insert a loxP-frt-neo-frt cassette upstream of exon 2, and a loxP site downstream of exon 4 of the dynactin 1 locus (Dctn1) on chromosome 6. The targeting vector was electroporated into 129/SvJ-derived embryonic stem cells. Chimeric males were bred to C57BL/6J females for germline transmission of the Dctn1^{LoxP-Frt-Neo-Frt} allele. The Dctn1^{LoxP-Frt-Neo-Frt} mice were bred to Rosa26^FlpE mice (on 129S4 genetic background; from Stock No. 003946) for germline deletion of the neo cassette - resulting in the Dctn1^LoxP/ (floxed exons 2-4) allele. The Dctn1^LoxP/ colony was further bred with transgenic strains expressing Thy1-Cre (on FVB/N genetic background; from Stock No. 006143) and/or CAG-Cre/Esr1 (on C57BL/6J genetic background; from Stock No. 004682). Dctn1^LoxP/ mice on a mixed C57BL/6J;FVB/N;129 genetic background (and not harboring the FlpE-, Cre- or Cre/Esr1-expressing alleles) were sent to The Jackson Laboratory Repository in 2018. 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

Symbol: Dctn1^tm2.1Cai
Name: targeted mutation 2.1, Huaibin Cai
MGI accession ID: MGI:6196333
Generation method: Targeted
Attributes: Conditional ready (e.g. floxed); No functional change
Synonyms: Dctn1^LoxP/
Marker symbol: Dctn1
Marker name: dynactin 1
Marker synonyms: DAP-150; DP-150; Glued; HMND14; P135; p150; p150^glued
Chromosome: 6
Strain of origin: 129X1/SvJ
Molecular note: A targeting vector was designed to insert a loxP-frt-neo-frt cassette upstream of exon 2, and a loxP site downstream of exon 4 of the dynactin 1 locus (Dctn1) on chromosome 6. Flp-mediated recombination removed the selection cassette.