Dual regulation of Stat1 and Stat3 by the tumor suppressor protein PML contributes to interferon α-mediated inhibition of angiogenesis

J Biol Chem. 2017 Jun 16;292(24):10048-10060. doi: 10.1074/jbc.M116.771071. Epub 2017 Apr 21.

Abstract

IFNs are effective in inhibiting angiogenesis in preclinical models and in treating several angioproliferative disorders. However, the detailed mechanisms of IFNα-mediated anti-angiogenesis are not completely understood. Stat1/2/3 and PML are IFNα downstream effectors and are pivotal regulators of angiogenesis. Here, we investigated PML's role in the regulation of Stat1/2/3 activity. In Pml knock-out (KO) mice, ablation of Pml largely reduces IFNα angiostatic ability in Matrigel plug assays. This suggested an essential role for PML in IFNα's anti-angiogenic function. We also demonstrated that PML shared a large cohort of regulatory genes with Stat1 and Stat3, indicating an important role of PML in regulating Stat1 and Stat3 activity. Using molecular tools and primary endothelial cells, we demonstrated that PML positively regulates Stat1 and Stat2 isgylation, a ubiquitination-like protein modification. Accordingly, manipulation of the isgylation system by knocking down USP18 altered IFNα-PML axis-mediated inhibition of endothelial cell migration and network formation. Furthermore, PML promotes turnover of nuclear Stat3, and knockdown of PML mitigates the effect of LLL12, a selective Stat3 inhibitor, on IFNα-mediated anti-angiogenic activity. Taken together, we elucidated an unappreciated mechanism in which PML, an IFNα-inducible effector, possess potent angiostatic activity, doing so in part by forming a positive feedforward loop with Stat1/2 and a negative feedback loop with Stat3. The interplay between PML, Stat1/Stat2, and Stat3 contributes to IFNα-mediated inhibition of angiogenesis, and disruption of this network results in aberrant IFNα signaling and altered angiostatic activity.

Keywords: STAT transcription factor; STAT3; angiogenesis; interferon; signal transducers and activators of transcription 1 (STAT1).

Publication types

  • Comparative Study

MeSH terms

  • Animals
  • Cell Line
  • Cells, Cultured
  • Endopeptidases / chemistry
  • Endopeptidases / genetics
  • Endopeptidases / metabolism
  • Endothelium, Vascular / cytology
  • Endothelium, Vascular / metabolism*
  • Endothelium, Vascular / pathology
  • Human Umbilical Vein Endothelial Cells / cytology
  • Human Umbilical Vein Endothelial Cells / metabolism
  • Humans
  • Interferon-alpha / genetics
  • Interferon-alpha / metabolism*
  • Mice, Knockout
  • Neovascularization, Pathologic / metabolism
  • Neovascularization, Pathologic / pathology
  • Neovascularization, Pathologic / prevention & control*
  • Neovascularization, Physiologic
  • Promyelocytic Leukemia Protein / antagonists & inhibitors
  • Promyelocytic Leukemia Protein / genetics
  • Promyelocytic Leukemia Protein / metabolism*
  • Protein Processing, Post-Translational
  • RNA Interference
  • Recombinant Proteins / metabolism
  • STAT1 Transcription Factor / agonists*
  • STAT1 Transcription Factor / genetics
  • STAT1 Transcription Factor / metabolism
  • STAT2 Transcription Factor / agonists*
  • STAT2 Transcription Factor / genetics
  • STAT2 Transcription Factor / metabolism
  • STAT3 Transcription Factor / antagonists & inhibitors*
  • STAT3 Transcription Factor / genetics
  • STAT3 Transcription Factor / metabolism
  • Ubiquitin Thiolesterase / antagonists & inhibitors
  • Ubiquitin Thiolesterase / genetics
  • Ubiquitin Thiolesterase / metabolism

Substances

  • Interferon-alpha
  • Pml protein, mouse
  • Promyelocytic Leukemia Protein
  • Recombinant Proteins
  • STAT1 Transcription Factor
  • STAT1 protein, human
  • STAT2 Transcription Factor
  • STAT2 protein, human
  • STAT3 Transcription Factor
  • STAT3 protein, human
  • Stat1 protein, mouse
  • Stat2 protein, mouse
  • Stat3 protein, mouse
  • PML protein, human
  • Endopeptidases
  • Usp18 protein, mouse
  • USP18 protein, human
  • Ubiquitin Thiolesterase