Expression of the tumour suppressor p53, often referred to as the “guardian of the genome”, is tightly regulated. However, critical details regarding the regulation and functions of p53 remain elusive [1]. It is known that the main negative regulator of p53 is the E3 ligase MDM2 that sequesters p53 and can ubiquitylate it for proteosomal degradation, keeping p53 protein levels low [2]. This regulation of p53 by MDM2 is frequently perturbed in a variety of cancers – often through MDM2 overexpression/amplification [3]. In response to therapeutic or oncogenic stressors, p53 becomes activated and transcriptionally upregulates a plethora of target genes controlling cellular pathways critical for tumour suppression such as cell death and growth arrest. One of the downstream p53 targets is the mdm2 gene itself, allowing for a negative feedback loop to control p53 expression[1, 4]. The relative importance of this feedback pathway in tumour suppression is not well understood.
To study the transcriptional regulation of mdm2, our lab has developed a novel reporter mouse model. Specifically, an IRES and the fluorophore Katushka-S2 coding sequence have been knocked-in downstream of the endogenous mdm2 gene. Thus, we can track changes in mdm2 expression in steady state and after p53 activation in various primary cell types.
Using this mdm2-Katushka-S2 reporter mouse, we have characterized mdm2 expression in a range of primary cell types ex vivo in the absence and presence of various p53-activating stimuli by flow cytometry. Additionally, we have performed intravital imaging of the bone marrow calvarium of the mdm2-Katushka-S2 reporter mice to assess mdm2 expression in response to g-irradiation in vivo. This reporter mouse model can further be used to assess how the MDM2-p53 negative feedback loop is perturbed in an oncogenic background and in response to cancer therapeutics, thereby allowing for a deeper understanding of p53 regulation and how the MDM2-p53 axis can best be targeted for therapy.