Generation and Characterization of Vav2 Genetically Engineered Mice

The methods used for the generation of Vav2^Onc/Onc knock-in (C57BL/6 background), Vav2^L332A/L332A knock-in (mixed genetic background), and Vav2^−/− knockout (C57BL/10 genetic background) mice have been described in previous publications^{37 (link),41 (link),76 (link)}. These mice and genetic background-matched WT controls were housed with an artificial 12-h light /12-h dark cycle under controlled temperature (23 °C) and humidity (50%) conditions. They were routinely maintained under ad libitum access to a standard chow global diet (Cat. #2018; Teklad global 18% protein) and tap water. When indicated, 8-week-old mice were shifted to either an HFD (45% fat, 4.73 kcal g⁻¹; Cat. #D12451; Research Diets) or a metformin regimen (2 mg mL⁻¹ in the drinking water; Cat. #PHR1084, Sigma-Aldrich) for 4 months before being euthanized. Endoplasmic reticulum stress was induced in the liver by injecting tunicamycin (2 μg kg⁻¹; Cat. #T7765, Sigma-Aldrich) intraperitoneally to 2-month-old mice. When indicated, 8-week-old mice were shifted to a methionine- and choline-deficient diet (Cat. #A02082002B, Research Diets) for the indicated period of time before being sacrificed. The experiments involving the in vivo infusion of insulin and IGF1 were performed indistinctly in age-matched male and female animals. The rest of the experiments were done using males.