A novel intracellular isoform of VEGFR‐1 activates Src and promotes cell invasion in MDA‐MB‐231 breast cancer cells

Abstract

Two types of VEGFR-1 receptors have been characterized: a full-length transmembrane receptor and a truncated extracellular soluble isoform (sVEGFR-1). We report here the characterization, in normal and cancer cells, of a new family of intracellular isoforms of VEGFR-1 resulting from alternative initiation of transcription in intronic sequences of the gene. While the classical isoforms of VEGFR-1 were barely detectable in MDA-MB-231 breast cancer cells, one of the intracellular isoforms transcribed from intron 21 (i(21)VEGFR-1) was the main isoform expressed in these cells. The new transcript encodes for a protein that contains only the phosphotransferase domain and the carboxyterminal tail of VEGFR-1. Treatment of MDA-MB-231 cells with siRNA specific for the tyrosine domain of VEGFR-1 suppressed the expression of i(21)VEGFR-1, downregulated phosphorylation of Src at tyrosine 418, and reduced markedly the invasion capacity of these cells in vitro. Accordingly, overexpression of transfected i(21)VEGFR-1 in MDA-MB-231 cells upregulated the active form of Src and increased invasiveness of MDA-MB-231 cells. The expression of i(21)VEGFR-1 in MDA-MB-231 cells was inhibited by retinoic acid. Both, activation of Src and downregulation by retinoic acid, have been reported in other intracellular members of the Fms/Kit/PDGFR family of tyrosine kinases, particularly in the intracellular isoform of c-kit, analogous structurally to i(21)VEGFR-1 and frequently expressed in cancer cells.

New pyrazolo[3,4-d]pyrimidine SRC inhibitors induce apoptosis in mesothelioma cell lines through p27 nuclear stabilization

“…The membranes were incubated with antibodies against phospho-SRC (raised in rabbit to phosphorylated Tyr416, which corresponds to human Tyr419) and SRC (Cell Signaling, Danvers, MA, USA), following blocking in 3% BSA. The double bands observed in some cell lines can derive from phospho-SRC antibody cross-reaction with different members of the SRC family kinases, as previously suggested (Mezquita et al, 2010) and consistently to what reported in MM specimens (Menges et al, 2010). The anti-b-actin antibody (Santa Cruz Biotechnology, Santa Cruz, CA, USA) was used for a loading control.…”

Tumour Cells Expressing Single VEGF Isoforms Display Distinct Growth, Survival and Migration Characteristics

“…The fibrosarcoma cells also expressed full length VEGFR1/flt1 as well as a truncated variant most likely consisting of the intracellular domain of the receptor (see supplementary data, Figure S2) previously described in endothelial and breast cancer cells [38]. The truncated variant was constitutively phosphorylated at tyr1333 in the fibrosarcomas cells but this phosphorylation could not be downregulated by SU11248.…”

Protein kinase C regulates FLT1 abundance and stimulates its cleavage in vascular endothelial cells with the release of a soluble PlGF/VEGF antagonist

“…sFLT1 isoforms, i13 and e15a, are encoded by transcripts that arise by intronic polyadenylation or by alternate splicing and result in a secreted protein that lacks the membrane-spanning and C-terminal domains (8, 9). Other transcripts that arise from FLT1 have intronic transcription start sties and encode isoforms that just contain portions of the intracellular tyrosine kinase domains and C-terminal tail (10). sFLT1 effectively functions as a natural VEGF antagonist regulating the actions of VEGF locally where it is produced and also in distant sites.…”