B6.Cg-Col1a1^{tm3(tetO-Mirlet7g/Mir21)Gqda}/J

Stock number: 023912
Research areas: Developmental Biology Research, Diabetes and Obesity Research

Description

These mice express Let-7S21L (let-7g (Mirlet7g) Stem, mir-21 (Mir21) Loop) under the regulatory control of a Col1a1 (collagen, type I, alpha 1)-driven tetO promoter and are useful in studies of growth and metabolism.

Detailed description

Lin28a (lin-28 homolog A (C. elegans)) and Lin28b (lin-28 homolog B (C. elegans)) encode RNA-binding proteins that block the biogenesis of Mirlet7 (let-7) microRNAs. Let-7 targets are known regulators of mammalian body size and metabolism.

This strain was constructed to determine whether altered let-7 expression might produce phenotypes opposite those of strains over-expressing Lin28a (see Stock No. 023910) or Lin28b (see Stock No. 0023911). It expresses Let-7S21L (let-7g Stem, mir-21 Loop) under the regulatory control of a Col1a1 (collagen, type I, alpha 1)-driven tetO promoter. Let-7S21L represents a chimeric Mirlet7g (microRNA let7g) stem/Mir21 (microRNA 21) loop fusion that cannot bind LIN28, thus allowing for let-7 processing despite Lin28 expression. This ensures that endogenous LIN28 cannot block pri- or pre-let-7g biosynthesis.

Mirlet7g/Mir21 expression can be regulated in progeny derived from crosses with rtTA or tTA mice. When crossed with Gt(ROSA)26Sor M2-rtTA strain (see Stock No. 006965), uninduced male mice exhibit no leakiness of expression and exhibit no growth or glucose phenotypes. Widespread induction from three weeks of age onward increases mature MIRLET7G levels in liver (>50-fold), skin (>20-fold), fat (approximately 4-fold) and muscle (approximately 4-fold). Induced animals show reduced body size and growth rates. After 5 days of induction, mice produce an increase in fed state glucose. Glucose tolerance testing reveals glucose intolerance in mice fed either a normal or high fat diet. Insulin tolerance testing fails to detect a difference in insulin sensitivity. Mutant mice produce more insulin than control mice.

Development

A targeting vector containing a splice acceptor, 2 polyadenylation sequences, a tetracycline operator, let-7S21L, and an SV40 polyadenylation signal within 3'UTR of the mouse Col1a1 gene was introduced to (C57BL/6 x 129S4/SvJae)F1-derived v6.5 embryonic stem (ES) cells containing M2-rtTA (optimized rtTA) targeted to the Gt(ROSA)26Sor locus promoter (see Stock No. 006965 for mutation details). Let-7S21L (let-7g Stem, mir-21 Loop) represents a chimeric Mirlet7g (microRNA let7g) stem/Mir21 (microRNA 21) loop fusion. Resultant chimeric males were crossed with outbred CD1 females. The Gt(ROSA)26Sor M2-rtTA mutation was bred away from this strain. The line was backcrossed to C57BL/6 for 8 generations by the donating lab.

Allele

Symbol: Col1a1^{tm3(tetO-Mirlet7g/Mir21)Gqda}
Name: targeted mutation 3, George Q Daley
MGI accession ID: MGI:5294613
Generation method: Targeted
Attributes: Inducible; Inserted expressed sequence; RMCE-ready
Marker symbol: Col1a1
Marker name: collagen, type I, alpha 1
Chromosome: 11
Strain of origin: (C57BL/6 x 129S4/SvJae)F1
Molecular note: RMCE on KH2 cells inserted a cassette that contains a splice acceptor, 2 polyadenylation sequence, tetracycline operator, a fusion of the Mirlet7g stem sequence and the Mir21 loop sequence, and an SV40 polyadenylation signal into the 3'UTR. No expression of the inserted sequence is detected in the absence of doxycycline treatment.
General note: The allele was made in KH2 cells that carry Gt(ROSA)26Sor^{tm1(rtTA*M2)Jae} and Col1a1^{tm13(neo/hygro*)Jae} (J:159351). This allele was generated with Col1a1^{tm13(neo/hygro*)Jae}.