Modelling Negative Feedback Networks for Activating Transcription Factor 3 Predicts a Dominant Role for miRNAs in Immediate Early Gene Regulation

Activating transcription factor 3 (Atf3) is rapidly and transiently upregulated in numerous systems, and is associated with various disease states. Atf3 is required for negative feedback regulation of other genes, but is itself subject to negative feedback regulation possibly by autorepression. In cardiomyocytes, Atf3 and Egr1 mRNAs are upregulated via ERK1/2 signalling and Atf3 suppresses Egr1 expression. We previously developed a mathematical model for the Atf3-Egr1 system. Here, we adjusted and extended the model to explore mechanisms of Atf3 feedback regulation. Introduction of an autorepressive loop for Atf3 tuned down its expression and inhibition of Egr1 was lost, demonstrating that negative feedback regulation of Atf3 by Atf3 itself is implausible in this context. Experimentally, signals downstream from ERK1/2 suppress Atf3 expression. Mathematical modelling indicated that this cannot occur by phosphorylation of pre-existing inhibitory transcriptional regulators because the time delay is too short. De novo synthesis of an inhibitory transcription factor (ITF) with a high affinity for the Atf3 promoter could suppress Atf3 expression, but (as with the Atf3 autorepression loop) inhibition of Egr1 was lost. Developing the model to include newly-synthesised miRNAs very efficiently terminated Atf3 protein expression and, with a 4-fold increase in the rate of degradation of mRNA from the mRNA/miRNA complex, profiles for Atf3 mRNA, Atf3 protein and Egr1 mRNA approximated to the experimental data. Combining the ITF model with that of the miRNA did not improve the profiles suggesting that miRNAs are likely to play a dominant role in switching off Atf3 expression post-induction.Author Summary: Activating transcription factor 3 (Atf3) is an important regulatory transcription factor which is associated with inflammation, restraint of the immune response and cancer. In this work, we develop a series of mathematical models to understand how Atf3 may be regulated. Informed with data from the literature and our own experiments, we show that self-regulation of Atf3 does not allow for variation between experimentally observed Atf3 mRNA and Atf3 protein expression profiles. A fast-acting signal via phosphorylated RSK is also shown to be implausible for similar reasons. Extending our mathematical model further, we postulate for the first time, that the observed dynamical variation in Atf3 mRNA and protein can be described by microRNAs downstream of RSKs. The further inclusion of an inhibitory transcription factor for Atf3 expression has little effect on these findings.