Sumoylation of Prox1 controls its ability to induce VEGFR3 expression and lymphatic phenotypes in endothelial cells.
Prox1 is a master regulator for the development of lymphatic vasculature and the induction of lymphangiogenesis. In this study, we identified Prox1 as a new target for small ubiquitin-like modifier 1 (SUMO-1). Lysine 556 (K556) was found to be the major sumoylation site for Prox1 in vitro and in vivo. Mutation of this site (from lysine to arginine K556R) reduced DNA binding and the transcriptional activity of Prox1. Overexpression of Prox1 in EA.hy926 endothelial cells induced expression of lymphatic endothelial cell-specific genes including vascular endothelial growth factor receptor 3 (VEGFR3), fibroblast growth factor receptor 3 (FGFR3) and p57 while expression of K556R mutant Prox1 had little effect. The induction of VEGFR3 by Prox1 in EA.hy926 endothelial cells was an indication of their response to VEGF-C-induced lymphangiogenic signals, including the enhancement of proliferation, sprouting and tube formation and the inhibition of apoptosis. This effect is SUMO-dependent because ectopic expression of SUMO-specific protease 2 (SENP2) effectively reduced Prox1 sumoylation and Prox1-induced VEGFR3 expression. In addition, K556R mutant Prox1 could not induce lymphatic phenotypes. Taken together, our results indicate that Prox1 is a target for SUMO-1 and suggest that sumoylation of Prox1 controls its ability to induce VEGFR3 expression and lymphatic phenotypes in endothelial cells.
Pan
Mei-Ren
MR
Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung 807, Taiwan.
Chang
Tsung-Ming
TM
Chang
Hui-Chiu
HC
Su
Jen-Liang
JL
Wang
Hsei-Wei
HW
Hung
Wen-Chun
WC
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D017209:Apoptosis; D002460:Cell Line; D049109:Cell Proliferation; D004247:DNA / Q000378:metabolism; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism*; D004729:Endothelium, Lymphatic / Q000166:cytology / Q000378:metabolism*; D018398:Homeodomain Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008239:Lysine / Q000378:metabolism; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000378:metabolism; D018919:Neovascularization, Physiologic; D010641:Phenotype; D011485:Protein Binding; D015398:Signal Transduction; D025841:Small Ubiquitin-Related Modifier Proteins / Q000378:metabolism*; D014158:Transcription, Genetic; D025521:Tumor Suppressor Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D042582:Vascular Endothelial Growth Factor C / Q000378:metabolism; D040321:Vascular Endothelial Growth Factor Receptor-3 / Q000378:metabolism*; D000098551:Prospero-Related Homeobox 1 Protein
D018398:Homeodomain Proteins; D050505:Mutant Proteins; D025841:Small Ubiquitin-Related Modifier Proteins; D025521:Tumor Suppressor Proteins; D042582:Vascular Endothelial Growth Factor C; D000098551:Prospero-Related Homeobox 1 Protein; D004247:DNA; D040321:Vascular Endothelial Growth Factor Receptor-3; D008239:Lysine