p19^Arf Represses Platelet-Derived Growth Factor Receptor β by Transcriptional and Posttranscriptional Mechanisms

Abstract

In addition to cancer surveillance, p19^Arf plays an essential role in blocking signals stemming from platelet-derived growth factor receptor β (Pdgfrβ) during eye development, but the underlying mechanisms have not been clear. We now show that without Arf, pericyte hyperplasia in the eye results from enhanced Pdgfrβ-dependent proliferation from embryonic day 13.5 (E13.5) of mouse development. Loss of Arf in the eye increases Pdgfrβ expression. In cultured fibroblasts and pericyte-like cells, ectopic p19^Arf represses and Arf knockdown enhances the expression of Pdgfrβ mRNA and protein. Ectopic Arf also represses primary Pdgfrβ transcripts and a plasmid driven by a minimal promoter, including one missing the CCAAT element required for high-level expression. p19^Arf uses both p53-dependent and -independent mechanisms to control Pdgfrβ. In vivo, without p53, Pdgfrβ mRNA is elevated and eye development abnormalities resemble the Arf ^−/− phenotype. However, effects of p53 on Pdgfrβ mRNA do not appear to be due to direct p53 or RNA polymerase II recruitment to the promoter. Although p19^Arf controls Pdgfrβ mRNA in a p53-dependent manner, it also blunts Pdgfrβ protein expression by blocking new protein synthesis in the absence of p53. Thus, our findings demonstrate a novel capacity for p19^Arf to control Pdgfrβ expression by p53-dependent and -independent mechanisms involving RNA transcription and protein synthesis, respectively, to promote the vascular remodeling needed for normal vision.

ACKNOWLEDGMENTS

We gratefully acknowledge C. Devitt (UT Southwestern) for technical assistance; L. A. Godley (University of Chicago) for providing C57BL/6 p53^+/− mice; S. Lowe (CSHL) for providing the shRNA-p53 construct; J. Sage (Stanford) for providing shRNA-p19^Arf constructs; C. J. Sherr (St. Jude Children's Research Hospital) for providing Arf^Gfp/Gfp mice; Y.-L. Chan, M. Pavon-Eternod, T. Pan, and I. Wool (all at the University of Chicago) for the use of equipment and for technical assistance with polysomal RNA isolation; G. P. Zambetti (St. Jude Children's Research Hospital) for providing TKO MEFs; and past and present members of the Skapek lab for helpful suggestions.

This work was supported by grants to S.X.S. from the National Institutes of Health (EY 014368 and EY 019942). C.Y.S. was supported by an institutional cardiology training grant (T32 HL007360-32).