Translationally controlled tumor protein (TCTP) is cytoplasmic and structurally related to guanine-nucleotide free chaperones. TCTP (also called histamine-releasing factor) has been described previously as a secreted protein that participates in inflammatory responses by promoting the release of histamine. How TCTP is eventually exported out of the cell to promote such activities is unknown. Here we show that TCTP secretion was insensitive to either brefeldin A or monensin, suggesting that it proceeds via an endoplasmic reticulum/Golgi-independent or nonclassical pathway. Moreover, our analyses also suggest that secreted TCTP originates from pre-existing pools. TSAP6, a p53-inducible 5-6 transmembrane protein, was found to interact with TCTP in a yeast two-hybrid hunt. GST pull down assays confirmed their direct interaction, and immunofluorescence analysis revealed their partial co-distribution to vesicular-like structures at the plasma membrane and around the nucleus. Functionally, the overexpression of TSAP6 consistently leads to enhanced secretion of both endogenously and exogenously expressed TCTP. Finally, we found TCTP in preparations of small secreted vesicles called exosomes, which have been suggested as a possible pathway for nonclassical secretion. Overexpression of TSAP6 also increased TCTP levels in exosome preparations. Altogether, these data identify a novel role for TSAP6 in the export of TCTP and indicate that this multipass membrane protein could have a general role in the regulation of vesicular trafficking and secretion.

Recently, we demonstrated that the expression levels of the translationally controlled tumor protein (TCTP) were strongly down-regulated at the mRNA and protein levels during tumor reversion͞suppression and by the activation of p53 and Siah-1. To better characterize the function of TCTP, a yeast two-hybrid hunt was performed. Subsequent analysis identified the translation elongation factor, eEF1A, and its guanine nucleotide exchange factor, eEF1B␤, as TCTP-interacting partners. In vitro and in vivo studies confirmed that TCTP bound specifically eEF1B␤ and eEF1A. Additionally, MS analysis also identified eEF1A as a TCTP interactor. Because eEF1A is a GTPase, we investigated the role of TCTP on the nucleotide exchange reaction of eEF1A. Our results show that TCTP preferentially stabilized the GDP form of eEF1A, and, furthermore, impaired the GDP exchange reaction promoted by eEF1B␤. These data suggest that TCTP has guanine nucleotide dissociation inhibitor activity, and, moreover, implicate TCTP in the elongation step of protein synthesis.

Neuropilin-1 and -2 (NRP1 and NRP2) are the transmembrane glycoproteins interacting with 2 types of ligands: class III semaphorins and several members of the VEGF family, the main regulators of blood and lymphatic vessel growth. We show here that both NRP1 and NRP2 can also bind hepatocyte growth factor (HGF). HGF is a pleiotropic cytokine and potent proangiogenic molecule that acts on its target cells by binding to the c-met receptor. We IntroductionNeuropilins (NRPs) are transmembrane glycoproteins that play an important role in various biological processes, including axonal guidance, angiogenesis, tumorigenesis, and the immunologic response. [1][2][3][4] NRPs have been characterized as coreceptors for 2 unrelated families of extracellular secreted ligands-class III semaphorins and several members of the vascular endothelial growth factor (VEGF) family, the main regulators of blood and lymphatic vessel growth. 5 NRP acts in conjunction with membraneassociated signal transducers, such as the VEGF receptor tyrosine kinases (VEGFR-1, -2, and -3) 6,7 and plexins, the transmembrane receptors of the semaphorin family. 8 In higher eukaryotes, 2 neuropilin genes, NRP1 and NRP2, have been identified. 9 They code for proteins displaying about 44% amino-acid sequence identity, with a similar domain structure. 10 Both NRP1 and NRP2 contain a large extracellular region and a short cytoplasmic tail of about 40 amino acids, lacking any enzymatic activity. The extracellular region of neuropilins contains 5 different structural domains-2 CUB motifs, a1 and a2, homologous to complement components C1r/C1s, 2 coagulation factor V/VIII homology domains b1 and b2, and one c domain (MAM, homologous to meprin, A5, ). 11 The high-affinity binding site for VEGF-A 165 has been localized to the b1 and b2 domains of NRP1 12,13 and NRP2, 14 whereas the binding of semaphorins requires both the a1a2 and b1b2 repeats. 12 NRP1 and NRP2 interact selectively with different members of the VEGF and semaphorin families and have nonoverlapping expression patterns. Thus, among the VEGF members, NRP1 binds VEGF-A 165 , VEGF-B, VEGF-E, and placental growth factor (PlGF), whereas NRP2 binds VEGF-A 165 ,VEGF-A 145 , VEGF-C, and PlGF. 15 The non-heparin-binding isoforms of VEGF, such as VEGF-A 121 , have long been considered unable to interact with NRPs. Current evidence suggests, however, that VEGF-A 121 does bind NRP1 via the C-terminal sequence of 6 amino acids encoded by exon 8. [16][17][18][19] During the development of the cardiovascular system, NRP1 is detected primarily in the arterial endothelial cells, and NRP2 is detected in the venous and lymphatic endothelial cells. 20 Genetic studies in mice have shown that both the overexpression of NRP1 21 and the targeted inactivation of the NRP1 gene 22,23 are lethal, provoking, in addition to neuronal defects, disorganization of the vascular network and defects in heart development. Inactivation of the NRP2 gene has less severe consequences, limited to defects in the formation of small lymphatic vessel...