Syndecan-syntenin-ALIX regulates the biogenesis of exosomes.

The biogenesis of exosomes, small secreted vesicles involved in signalling processes, remains incompletely understood. Here, we report evidence that the syndecan heparan sulphate proteoglycans and their cytoplasmic adaptor syntenin control the formation of exosomes. Syntenin interacts directly with ALIX through LYPX(n)L motifs, similarly to retroviral proteins, and supports the intraluminal budding of endosomal membranes. Syntenin exosomes depend on the availability of heparan sulphate, syndecans, ALIX and ESCRTs, and impact on the trafficking and confinement of FGF signals. This study identifies a key role for syndecan-syntenin-ALIX in membrane transport and signalling processes.

Syntenin, a syndecan adaptor and an Arf6 phosphatidylinositol 4,5-bisphosphate effector, is essential for epiboly and gastrulation cell movements in zebrafish.

Epiboly, the spreading and the thinning of the blastoderm to cover the yolk cell and close the blastopore in fish embryos, is central to the process of gastrulation. Despite its fundamental importance, little is known about the molecular mechanisms that control this coordinated cell movement. By a combination of knockdown studies and rescue experiments in zebrafish (Danio rerio), we show that epiboly relies on the molecular networking of syntenin with syndecan heparan sulphate proteoglycans, which act as co-receptors for adhesion molecules and growth factors. Furthermore, we show that the interaction of syntenin with phosphatidylinositol 4,5-bisphosphate (PIP2) and with the small GTPase ADP-ribosylation factor 6 (Arf6), which regulate the endocytic recycling of syndecan, is necessary for epiboly progression. Analysis of the earliest cellular defects suggests a role for syntenin in the autonomous vegetal expansion of the yolk syncytial layer and the rearrangement of the actin cytoskeleton in extra-embryonic tissues, but not in embryonic cell fate determination. This study identifies the importance of the syntenin-syndecan-PIP2-Arf6 complex for the progression of fish epiboly and establishes its key role in directional cell movements during early development.

The PDZ2 domain of zonula occludens-1 and -2 is a phosphoinositide binding domain.

Zonula occludens proteins (ZO) are postsynaptic density protein-95 discs large-zonula occludens (PDZ) domain-containing proteins that play a fundamental role in the assembly of tight junctions and establishment of cell polarity. Here, we show that the second PDZ domain of ZO-1 and ZO-2 binds phosphoinositides (PtdInsP) and we identified critical residues involved in the interaction. Furthermore, peptide and PtdInsP binding of ZO PDZ2 domains are mutually exclusive. Although lipid binding does not seem to be required for plasma membrane localisation of ZO-1, phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P (2)) binding to the PDZ2 domain of ZO-2 regulates ZO-2 recruitment to nuclear speckles. Knockdown of ZO-2 expression disrupts speckle morphology, indicating that ZO-2 might play an active role in formation and stabilisation of these subnuclear structures. This study shows for the first time that ZO isoforms bind PtdInsPs and offers an alternative regulatory mechanism for the formation and stabilisation of protein complexes in the nucleus.

The postsynaptic density 95/disc-large/zona occludens protein syntenin directly interacts with frizzled 7 and supports noncanonical Wnt signaling.

Wnt signaling pathways are essential for embryonic patterning, and they are disturbed in a wide spectrum of diseases, including cancer. An unresolved question is how the different Wnt pathways are supported and regulated. We previously established that the postsynaptic density 95/disc-large/zona occludens (PDZ) protein syntenin binds to syndecans, Wnt coreceptors, and known stimulators of protein kinase C (PKC)alpha and CDC42 activity. Here, we show that syntenin also interacts with the C-terminal PDZ binding motif of several Frizzled Wnt receptors, without compromising the recruitment of Dishevelled, a key downstream Wnt-signaling component. Syntenin is coexpressed with cognate Frizzled during early development in Xenopus. Overexpression and down-regulation of syntenin disrupt convergent extension movements, supporting a role for syntenin in noncanonical Wnt signaling. Syntenin stimulates c-jun phosphorylation and modulates Frizzled 7 signaling, in particular the PKCalpha/CDC42 noncanonical Wnt signaling cascade. The syntenin-Frizzled 7 binding mode indicates syntenin can accommodate Frizzled 7-syndecan complexes. We propose that syntenin is a novel component of the Wnt signal transduction cascade and that it might function as a direct intracellular link between Frizzled and syndecans.

Syndecan-3 is a dendritic cell-specific attachment receptor for HIV-1.

Dendritic cells (DCs) efficiently capture HIV-1 and mediate transmission to T cells, but the underlying molecular mechanism is still being debated. The C-type lectin DC-SIGN is important in HIV-1 transmission by DCs. However, various studies strongly suggest that another HIV-1 receptor on DCs is involved in the capture of HIV-1. Here we have identified syndecan-3 as a major HIV-1 attachment receptor on DCs. Syndecan-3 is a DC-specific heparan sulfate (HS) proteoglycan that captures HIV-1 through interaction with the HIV-1 envelope glycoprotein gp120. Syndecan-3 stabilizes the captured virus, enhances DC infection in cis, and promotes transmission to T cells. Removal of the HSs from the cell surface by heparinase III or by silencing syndecan-3 by siRNA partially inhibited HIV-1 transmission by immature DCs, whereas neutralizing both syndecan-3 and DC-SIGN completely abrogated HIV-1 capture and subsequent transmission. Thus, HIV-1 exploits both syndecan-3 and DC-SIGN to mediate HIV-1 transmission, and an effective microbicide should target both syndecan-3 and DC-SIGN on DCs to prevent transmission.

Syndecan recycling [corrected] is controlled by syntenin-PIP2 interaction and Arf6.

Syndecans are heparan sulfate proteoglycans that modulate the activity of several growth factors and cell adhesion molecules. PDZ domains in the adaptor protein syntenin interact with syndecans and with the phosphoinositide PIP(2), which is involved in the regulation of the actin cytoskeleton and membrane trafficking. Here, we show that the syntenin PDZ domain-PIP(2) interaction controls Arf6-mediated syndecan recycling through endosomal compartments. FGF receptor accompanies syndecan along the syntenin-mediated recycling pathway, in a heparan sulfate- and FGF-dependent manner. Syndecans that cannot recycle via this pathway become trapped intracellularly and inhibit cell spreading. This syntenin-mediated syndecan recycling pathway may regulate the surface availability of a number of cell adhesion and signaling molecules.

Nuclear speckles and nucleoli targeting by PIP2-PDZ domain interactions.

PDZ (Postsynaptic density protein, Disc large, Zona occludens) domains are protein-protein interaction modules that predominate in submembranous scaffolding proteins. Recently, we showed that the PDZ domains of syntenin-1 also interact with phosphatidylinositol 4,5-bisphosphate (PIP2) and that this interaction controls the recruitment of the protein to the plasma membrane. Here we evaluate the general importance of PIP2-PDZ domain interactions. We report that most PDZ proteins bind weakly to PIP2, but that syntenin-2, the closest homolog of syntenin-1, binds with high affinity to PIP2 via its PDZ domains. Surprisingly, these domains target syntenin-2 to nuclear PIP2 pools, in nuclear speckles and nucleoli. Targeting to these sites is abolished by treatments known to affect these PIP2 pools. Mutational and domain-swapping experiments indicate that high-affinity binding to PIP2 requires both PDZ domains of syntenin-2, but that its first PDZ domain contains the nuclear PIP2 targeting determinants. Depletion of syntenin-2 disrupts the nuclear speckles-PIP2 pattern and affects cell survival and cell division. These findings show that PIP2-PDZ domain interactions can directly contribute to subnuclear assembly processes.

Processing by proprotein convertases is required for glypican-3 modulation of cell survival, Wnt signaling, and gastrulation movements.

Glypican (GPC)-3 inhibits cell proliferation and regulates cell survival during development. This action is demonstrated by GPC3 loss-of-function mutations in humans and mice. Here, we show that the GPC3 core protein is processed by a furinlike convertase. This processing is essential for GPC3 modulating Wnt signaling and cell survival in vitro and for supporting embryonic cell movements in zebrafish. The processed GPC3 core protein is necessary and sufficient for the cell-specific induction of apoptosis, but in vitro effects on canonical and noncanonical Wnt signaling additionally require substitution of the core protein with heparan sulfate. Wnt 5A physically associates only with processed GPC3, and only a form of GPC3 that can be processed by a convertase is able to rescue epiboly and convergence/extension movements in GPC3 morphant embryos. Our data imply that the Simpson-Golabi-Behmel syndrome may in part result from a loss of GPC3 controls on Wnt signaling, and suggest that this function requires the cooperation of both the protein and the heparan sulfate moieties of the proteoglycan.