Semaphorin 3d and Semaphorin 3e Direct Endothelial Motility through Distinct Molecular Signaling Pathways

• Published in: The Journal of biological chemistry 2014-06, Vol.289 (26), p.17971-17979
• Main Authors: Aghajanian, Haig, Choi, Connie, Ho, Vivienne C., Gupta, Mudit, Singh, Manvendra K., Epstein, Jonathan A.
• Format: Article
• Language: English
• Subjects: Actin; Akt; Animals; Cell Adhesion Molecules, Neuronal - genetics; Cell Adhesion Molecules, Neuronal - metabolism; Cell Motility; Cell Movement; Cytoskeletal Proteins; Cytoskeleton - genetics; Cytoskeleton - metabolism; Developmental Biology; Endothelial Cell; Endothelial Cells - cytology; Endothelial Cells - enzymology; Endothelial Cells - metabolism; Female; Glycoproteins - genetics; Glycoproteins - metabolism; HEK293 Cells; Humans; Intracellular Signaling Peptides and Proteins; Male; Membrane Glycoproteins; Membrane Proteins - genetics; Membrane Proteins - metabolism; Mice; Mice, Knockout; Nerve Tissue Proteins; Neuropilin; Neuropilin-1 - genetics; Neuropilin-1 - metabolism; Phosphatidylinositol 3-Kinases - genetics; Phosphatidylinositol 3-Kinases - metabolism; PI3K; Plexin; Proto-Oncogene Proteins c-akt - genetics; Proto-Oncogene Proteins c-akt - metabolism; Semaphorin; Semaphorins - genetics; Semaphorins - metabolism; Signal Transduction; Smad3 Protein - genetics; Smad3 Protein - metabolism
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M113.544833

Class 3 semaphorins were initially described as axonal growth cone guidance molecules that signal through plexin and neuropilin coreceptors and since then have been established to be regulators of vascular development. Semaphorin 3e (Sema3e) has been shown previously to repel endothelial cells and is the only class 3 semaphorin known to be capable of signaling via a plexin receptor without a neuropilin coreceptor. Sema3e signals through plexin D1 (Plxnd1) to regulate vascular patterning by modulating the cytoskeleton and focal adhesion structures. We showed recently that semaphorin 3d (Sema3d) mediates endothelial cell repulsion and pulmonary vein patterning during embryogenesis. Here we show that Sema3d and Sema3e affect human umbilical vein endothelial cells similarly but through distinct molecular signaling pathways. Time-lapse imaging studies show that both Sema3d and Sema3e can inhibit cell motility and migration, and tube formation assays indicate that both can impede tubulogenesis. Endothelial cells incubated with either Sema3d or Sema3e demonstrate a loss of actin stress fibers and focal adhesions. However, the addition of neuropilin 1 (Nrp1)-blocking antibody or siRNA knockdown of Nrp1 inhibits Sema3d-mediated, but not Sema3e-mediated, cytoskeletal reorganization, and siRNA knockdown of Nrp1 abrogates Sema3d-mediated, but not Sema3e-mediated, inhibition of tubulogenesis. On the other hand, endothelial cells deficient in Plxnd1 are resistant to endothelial repulsion mediated by Sema3e but not Sema3d. Unlike Sema3e, Sema3d incubation results in phosphorylation of Akt in human umbilical vein endothelial cells, and inhibition of the PI3K/Akt pathway blocks the endothelial guidance and cytoskeletal reorganization functions of Sema3d but not Sema3e. Background: Class 3 semaphorins are guidance molecules for endothelial cells. Results: In multiple endothelial cell assays, semaphorin 3d requires neuropilin 1 or PI3K/Akt but not plexin D1, whereas semaphorin 3e requires plexin D1 but not neuropilin 1 or PI3K/Akt. Conclusion: Semaphorin 3d and 3e utilize different pathways to mediate similar effects in endothelial cells. Significance: Related guidance molecules utilize distinct mechanisms to repel endothelial cells.