Development of shRNA transgenic mice for preclinical studies of the MKK4 regeneration kinase

Abstract

The liver harbors a distinct capacity for endogenous regeneration; however, liver regeneration is often impaired in disease and therefore insufficient to compensate for the loss of hepatocytes and organ function. We recently identified MKK4 as a key regulator of liver regeneration by an in vivo RNAi screen. We could show that the knock down of MKK4 in hepatocytes increases their regenerative capacity and robustness. For further investigations we set out to generate shRNA transgenic mouse lines for inducible and reversible gene silencing of MKK4. We used mir30 based MKK4-shRNAs and placed them in the 3' untranslated region of the fluorescent reporter GFP controlled by a tet-responsive element (TREt), generating a TRE based Col1a1 targeting vector. Generated mice were then crossed to the CAGs-rtTA3 mouse line, which expresses the reverse tet-transactivator ubiquitously. The application of the tetracycline analog doxycycline allowed for a regulated expression from the TRE promoter and thereby expression of the MKK4 shRNA. Using these newly generated mouse lines we are able to study phenotypes after an ubiquitous MKK4 silencing in vivo. Current studies are aiming to analyze the role of MKK4 as a potential regeneration kinase in tissues other than liver.