|Ph.D Student||Rabinowicz Noa|
|Subject||The Role of Neuropilin-1 and Plexin-A4 in Angiogenesis|
and Tumor Progression
|Department||Department of Medicine||Supervisor||Professor Emeritus Gera Neufeld|
|Full Thesis text|
Neuropilin-1 (Np1) functions as a receptor for heparin binding forms of vascular endothelial growth factor (VEGF) such as VEGF165. A shorter VEGF isoform, VEGF121, does not bind Np1. Despite that, Np1 functions as an enhancer that potentiates both VEGF165 and VEGF121 signaling, via the VEGFR-2 tyrosine-kinase receptor. To study the mechanism by which neuropilin-1 potentiates VEGF activity we produced a mutant neuropilin-1 (Np13mut) that doesn't bind VEGF165. Expression of this mutant in endothelial cells enhanced VEGF-induced VEGFR-2 phosphorylation, as did wild-type Np1. However, while wt-Np1 formed increased amount of complexes with VEGFR-2 upon exposure to VEGF165, Np13mut did not, suggesting that the VEGF binding site of Np1 is dispensable for Np1 in order to enhance VEGF signaling through VEGFR-2, and that basal Np1-VEGFR-2 complexes are enough for this enhancement.
Np1 is also known to promote tumor progression, and in some cancers its expression correlates to poor prognosis. Upon its expression in U87MG glioblastoma cells, Np13mut increased tumor cell proliferation in-vitro and tumor growth in-vivo, as did wt-Np1. The tumor vasculature was similar in control and Np1 expressing cells, suggesting that Np1 affected the tumor cells directly and not via an effect on the surrounding endothelial cells.
Another group of receptors which are expressed on endothelial cells are the Plexins, semaphorins receptors. Plexin-A4 is a receptor for sema6A and sema6B and associates with neuropilins to transduce signals of class-3 semaphorins. We observed that plexin-A1 and plexin-A4 are required simultaneously for transduction of inhibitory sema3A signals and that they form complexes. Inhibition of plexin-A1 or plexin-A4 expression in endothelial cells using specific shRNAs resulted in prominent plexin type specific rearrangements of the actin cytoskeleton that were accompanied by inhibition of bFGF and VEGF induced cell proliferation. The two responses were not interdependent since silencing plexin-A4 in U87MG glioblastoma cells inhibited cell proliferation and the formation of tumors from these cells without affecting cytoskeletal organization. Plexin-A4 formed complexes with FGFR1 and VEGFR-2 and enhanced VEGF-induced VEGFR-2 phosphorylation in endothelial cells as well as bFGF-induced cell proliferation. Silencing plexin-A4 also resulted in the inhibition of angiogenesis in in-vitro assays.
Some of the pro-proliferative effects of plexin-A4 were due to transduction of autocrine sema6B induced pro-proliferative signals, since silencing sema6B expression in endothelial and U87MG cells mimicked the effects of plexin-A4 silencing and inhibited tumor formation of U87MG cells. These results suggest that plexin-A4 may represent a target for developing novel anti-angiogenic and anti-tumorigenic drugs.