A Complex Regulatory Network of Transcription Factors Critical for Ocular Development and Disease

The PITX2 \"homeobox\" and FOXC1 and FOXC2 \"forkhead box\" transcription factors are critical for eye development and cause human ocular diseases when mutated. We have identified biochemical and genetic links between these transcription factors and a transcriptional regulator protein PRKC apoptosis WT-1 regulator (PAWR) that we propose functionally connect all these proteins in a common pathway critically involved in eye development. We discovered all binary physical interactions between FOXC1, PITX2, FOXC2 and PAWR. Importantly, PAWR modulates the abilities of PITX2, FOXC1 and FOXC2 to activate their genetic targets. Together with either FOXC1 or FOXC2, PAWR increases PITX2 activity. PAWR reduces PITX2 activity in absence of FOXC1 or FOXC2. At the same time PAWR also exerts different regulatory effects on different FOXC target sites. Furthermore, morpholino knockdown of pitx2, foxc1 and pawr in zebrafish indicate that PAWR, FOXC1 and PITX2 genetically interact, and are in the same developmental pathway. These data for the first time tie PITX2, FOXC1, FOXC2 and PAWR into a common regulatory pathway. We have therefore identified a functional link between three transcription factors, modulated by PAWR, which we propose underlies the similar ocular phenotypes and glaucoma pathology caused by mutations of these genes.