Syndapin: Biological Overview | References
Gene name - Syndapin
Cytological map position - 93A1-93A2
Function - lipid-binding protein
Keywords - F-BAR-domain containing protein that regulates cellularization, membrane tubulation, Clathrin-mediated and bulk endocytosis, formation of subsynaptic reticulum and F-actin stability, contributes to coupling the plasma membrane and contractile ring in cytokinesis
Symbol - Synd
FlyBase ID: FBgn0053094
Genetic map position - chr3R:20,832,799-20,840,118
NCBI classification - F-BAR_PACSIN, SH3_PACSIN
Cellular location - lipid membranes
|Recent literature||Schultheis, D., Schwirz, J. and Frasch, M. (2019). RNAi screen in Tribolium reveals involvement of F-BAR proteins in myoblast fusion and visceral muscle morphogenesis in insects. G3 (Bethesda). PubMed ID: 30733382
In a large-scale RNAi screen in Tribolium castaneum for genes with knock-down phenotypes in the larval somatic musculature, one recurring phenotype was the appearance of larval muscle fibers that were significantly thinner than those in control animals. Several of the genes producing this knock-down phenotype corresponded to orthologs of Drosophila genes that are known to participate in myoblast fusion, particularly via their effects on actin polymerization. A new gene previously not implicated in myoblast fusion but displaying a similar thin-muscle knock-down phenotype was the Tribolium ortholog of Nostrin, which encodes an F-BAR and SH3 domain protein. Genetic studies of Nostrin and Cip4, a gene encoding a structurally related protein, in Drosophila show that the encoded F-BAR proteins jointly contribute to efficient myoblast fusion during larval muscle development. Together with the F-Bar protein Syndapin they are also required for normal embryonic midgut morphogenesis. In addition, Cip4 is required together with Nostrin during the profound remodeling of the midgut visceral musculature during metamorphosis. It is proposed that these F-Bar proteins help govern proper morphogenesis particularly of the longitudinal midgut muscles during metamorphosis.
|Ogi, S., Matsuda, A., Otsuka, Y., Liu, Z., Satoh, T. and Satoh, A. K. (2019). Syndapin constricts microvillar necks to form a united rhabdomere in Drosophila photoreceptors. Development. PubMed ID: 31371377
Drosophila photoreceptors develop from polarized epithelial cells that have apical and basolateral membranes. During morphogenesis, the apical membranes subdivide into a united bundle of photosensory microvilli (rhabdomeres) and a surrounding supporting membrane (stalk). By EMS-induced mutagenesis screening, this study found that the F-Bin/Amphiphysin/Rvs (F-BAR) protein Syndapin is essential for apical membrane segregation. The analysis of the super-resolution microscopy, STORM and the electron microscopy suggest that Syndapin localizes to the neck of the microvilli at the base of the rhabdomere. Syndapin and Moesin are required to constrict the neck of the microvilli to organize the membrane architecture at the base of the rhabdomere, to exclude the stalk membrane. Simultaneous loss of syndapin along with the microvilli adhesion molecule chaoptin significantly enhanced the disruption of stalk-rhabdomere segregation. However, loss of the factors involving endocytosis do not interfere. These results indicated Syndapin is most likely functioning through its membrane curvature properties, and not through endocytic processes for stalk-rhabdomere segregation. Elucidation of the mechanism of this unconventional domain formation will provide novel insights into the field of cell biology.
|Sherlekar, A., Mundhe, G., Richa, P., Dey, B., Sharma, S. and Rikhy, R. (2020). F-BAR domain protein Syndapin regulates actomyosin dynamics during apical cap remodeling in syncytial Drosophila embryos. J Cell Sci. PubMed ID: 32327556
Branched actin networks driven by Arp2/3 collaborate with actomyosin filaments in processes such as cell migration. The syncytial Drosophila blastoderm embryo also shows expansion of apical caps by Arp2/3 driven actin polymerization in interphase and buckling at contact edges by MyosinII to form furrows in metaphase. The role of Syndapin (Synd), an F-BAR domain containing protein in apical cap remodelling prior to furrow extension. synd depletion showed larger apical caps. STED super-resolution and TIRF microscopy showed long apical actin protrusions in caps in interphase and short protrusions in metaphase in control embryos. synd depletion led to sustained long protrusions even in metaphase. Loss of Arp2/3 function in synd mutants partly reverted defects in apical cap expansion and protrusion remodelling. MyosinII levels were decreased in synd mutants and MyosinII mutant embryos have been previously reported to have expanded caps. It is proposed that Syndapin function limits branching activity during cap expansion and affects MyosinII distribution in order to shift actin remodeling from apical cap expansion to favor lateral furrow extension.
Coordinated membrane and cytoskeletal remodeling activities are required for membrane extension in processes such as cytokinesis and syncytial nuclear division cycles in Drosophila. Pseudocleavage furrow membranes in the syncytial Drosophila blastoderm embryo show rapid extension and retraction regulated by actin-remodeling proteins. The F-BAR domain protein Syndapin (Synd) is involved in membrane tubulation, endocytosis, and, uniquely, in F-actin stability. This study reports a role for Synd in actin-regulated pseudocleavage furrow formation. Synd localized to these furrows, and its loss resulted in short, disorganized furrows. Synd presence was important for the recruitment of the septin Peanut and distribution of Diaphanous and F-actin at furrows. Synd and Peanut were both absent in furrow-initiation mutants of RhoGEF2 and Diaphanous and in furrow-progression mutants of Anillin. Synd overexpression in rhogef2 mutants reversed its furrow-extension phenotypes, Peanut and Diaphanous recruitment, and F-actin organization. It is concluded that Synd plays an important role in pseudocleavage furrow extension, and this role is also likely to be crucial in cleavage furrow formation during cell division (Sherlekar, 2016).
Cleavage furrow formation during cell division requires a highly conserved set of cytoskeletal and membrane-trafficking proteins. Their positioning and initiation involves microtubules and the centralspindlin complex. Rho-GTPase-activating protein (RacGAP50C) of this complex positions Rho-GTP exchange factor (RhoGEF) Pebble at contractile rings, and another RhoGEF2 functions in pseudocleavage furrows to activate Rho1 for furrow initiation. Rho1 recruits formins that assemble an actin scaffold for contractile-ring formation and/or furrow initiation. Formin activity also depends on the presence of a scaffold protein, Anillin, at the contractile ring. RacGAP50C also accumulates Anillin at the furrow, which is responsible for both septin and myosin II association at the contractile ring. Cytokinesis failure increases in Caenorhabditis elegans when embryos are depleted of both Rho kinase and Anillin/septins, implying that they work together for robust furrow formation (Sherlekar, 2016).
The cell division cycle is accompanied by drastic changes in cell shape that necessitate dynamic interplay between the membrane and actin cytoskeleton. In the Drosophila syncytial embryo, nuclear division cycles 10-13 are rapid and involve dynamic pseudocleavage furrow ingression and retraction between adjacent dividing nuclei. These furrows serve to prevent spindle cross-talk across compartments during metaphase of each cycle and organize the embryo into discrete polarized functional units. Furrow positioning and initiation at this stage requires RhoGEF2 for recruiting Rho1 and the formin Diaphanous (Dia). Microtubules are required for furrow positioning, while furrow ingression involves dynamic growth of actin filaments through Profilin and the action of anticapping proteins (like Ena/VASP). The syncytial cycles are followed by massive elongation of furrows to form individual cells in a process called cellularization, during which membrane extension is fueled by flattening of apical microvilli and Rab11-mediated endocytosis and driven by an actomyosin contractile ring that, apart from actin and myosin II, also comprises Anillin, septins, RhoGEF2, and Dia. Although contractile rings first form only during cellularization in early developing Drosophila embryos, the syncytial pseudocleavage furrows contain most of the proteins present in the contractile ring such as Rho1, RhoGEF2, Dia, Anillin, and septins (Sherlekar, 2016).
F-BAR domain-containing proteins link membrane and cytoskeleton in various processes, including endocytosis, cell shape and polarity, cell motility, and cytokinesis. The yeast orthologues of F-BAR protein Cip4 are known to recruit formins and influence their nucleation and elongation activities. In addition, Hof1 (Cip4 in Saccharomyces cerevisiae) coiled-coil domain binds Septin (Cdc10) and localizes it to the bud neck. Drosophila Cip4, however, is not essential for formin Dia recruitment to cellularization furrows, and its loss does not result in a defect in cellularization but its overexpression shows dia loss-of-function phenotypes. The F-BAR domain protein, Syndapin/Pacsin (Synd), initially identified as a binding partner for Dynamin and neuronal Wiscott-Aldrich syndrome protein (N-Wasp) via its SH3 domain, participates in endocytosis and actin remodeling (Roos, 1998; Simpson, 1999; Qualmann, 2000; Dharmalingam, 2009; Rao, 2010). Mammalian Synd1 binds to the actin nucleator Cordon bleu (Cobl) (Ahuja, 2007) and mediates its interaction with Arp2/3 to affect actin nucleation during neuromorphogenesis (Schwintzer, 2011). Synd, unlike other F-BAR proteins, directly binds and stabilizes F-actin (Kostan, 2014) and, unlike any N- or F-BAR protein, can generate a range of membrane curvatures much greater than its own intrinsic curvature (Frost, 2008; Ramesh, 2013). Drosophila Synd promotes expansion of the subsynaptic reticulum (Kumar, 2009b), which also requires actin-remodeling (Ramachandran, 2009). Drosophila Synd also binds to Anillin via its myosin-binding domain in vitro, localizes at the cytokinetic furrow (earlier than Drosophila Cip4) in D.Mel-2 cells, and is important for cytokinesis during male meiosis in primary spermatocytes (Takeda, 2013). Together these studies suggest a role for Synd in coordinated membrane and actin remodeling during cleavage furrow formation. However, no analysis of its recruitment dynamics or functional analysis in organization of actin or actin-remodeling proteins with respect to furrow initiation or extension machinery has been carried out so far. This study reports that Synd is important for syncytial Drosophila pseudocleavage furrow extension; septin Peanut (Pnut) recruitment; and distinct Dia, Anillin, and actin localization. Most significantly, Synd can recruit actin remodeling proteins, organize actin, and result in furrow extension during pseudocleavage furrow formation in rhogef2-depleted embryos (Sherlekar, 2016).
Syndapins belong to the family of highly conserved F-BAR-domain containing proteins with diverse roles in membrane tubulation, Clathrin-mediated and bulk endocytosis, and actin remodeling and cytokinesis. Synd is thus poised to play a role in processes like furrow formation, which needs orchestrated remodeling of both the membrane and the cytoskeleton. Furrow elongation in syncytial Drosophila embryos is an excellent model system to study the role of proteins that drive its formation. Previous studies show that furrow formation involves membrane addition by trafficking and membrane extension by remodeling of the actin meshwork. This study has conclusively demonstrated that Synd functions to promote furrow formation by organization and elongation of F-actin structures. Synd is essential for recruitment and distribution of Pnut and Dia on the membrane. In turn, Pnut and Dia also affect Synd distribution on the membrane. RhoGEF2/Dia and Anillin/Pnut have been previously shown to regulate F-actin architecture at cleavage and cellularization furrows, and loss of Synd in synd mutants therefore affects actin both directly and through its influence on Pnut and Dia localization. As with other actin-regulated processes, even though a linear pathway of association/regulation of these actin-remodeling proteins to the furrow membrane is unlikely, the data imply that Synd is a key component in the RhoGEF2-Dia-Anillin/Pnut pathway during actin-driven furrow elongation. Synd2 can bind and inhibit Rac1 via its SH3 domain, thus reducing Arp2/3 activity, and may therefore be able to potentiate Dia activity by increasing RhoA levels. Such a mechanism can explain increased Dia function when Synd is overexpressed in RhoGEF2 knockdown embryos, which, along with recruitment of Pnut to the membrane, can help organize actin and elongate cleavage furrows (Sherlekar, 2016).
Actin stabilization into continuous structures reversed the furrow length defect in synd mutant embryos. Jasplakinolide (Jasp) blocks actin turnover at the contractile ring and affects cleavage furrow invagination while preserving furrow integrity, and hence showed fewer punctae in synd and rhogef2 mutant embryos. CytoD, on the other hand, allows actin polymerization, and as a result, synd and rhogef2 mutant embryos treated with CytoD displayed more organized actin structures and elongated furrows. This provides mechanistic insight into how Synd functions in regulating actin polymerization and may be further investigated through kinetic studies of actin polymerization (Sherlekar, 2016).
Overexpression of Synd and not Pnut in the rhogef2RNAi-containing embryos partially reversed pseudocleavage furrow recruitment and morphology defects seen in rhogef2RNAi and increased the furrow length compared with wild type. Synd activity is thus needed at the pseudocleavage furrow for extension, and some as yet uncharacterized proteins play a role in furrow limitation. It is interesting to compare the functions of F-BAR domain proteins, Synd with Cip4 in furrow elongation and Dia recruitment. Cip4 antagonizes Dia function, and its overexpression has dia loss-of-function phenotypes like missing furrows. It is possible that opposing activities of F-BAR proteins Synd and Cip4 with respect to Dia are in a balance, and future experiments can test whether this function plays a role in limiting the growth of pseudocleavage furrows (Sherlekar, 2016).
Because Synd's SH3 domain interacts with Dynamin, and Dynamin has a role in endocytosis and furrow extension in syncytial divisions and cellularization, it remained to be investigated whether Clathrin-dependent endocytosis defects in synd mutants also affect furrow elongation. This study shows that synd mutant embryos have defects in cleavage furrow-tubule length and Rab5 vesicle numbers. Decrease in Rab5 vesicle numbers is also seen in rhogef2 mutant embryos. However, Synd-GFP overexpression in rhogef2 mutant embryos is able to reverse the furrow-extension defect without rescuing the Rab5 endocytic vesicle defect. Taken together, these data show that Synd has a role in endocytosis, but the reversal of furrow phenotypes in rhogef2 mutant embryos is due to the ability of Synd to recruit and organize actin and proteins of the actin-remodeling machinery such as Dia and Pnut (Sherlekar, 2016).
This analysis of membrane architecture and pseudocleavage furrow length in rhogef2, pnut, and synd mutants found that shorter furrows in each of these mutants were also loose/unstable and had slow lateral movement during the nuclear cycle. Septins brace the plasma membrane against aberrant cell-shape deformation and are able to tubulate phosphatidylinositol-4,5-bisphosphate liposome membranes when treated with a brain extract. It is probable that Septin-mediated membrane tubulation activity and cell-shape effects are dependent on the presence of F-BAR proteins like Synd. Sept7 mutants in Xenopus show unstable and undulating membranes during gastrulation. This substantiates Synd’s role in maintenance of membrane integrity and shape by affecting actin organization and Pnut recruitment (Sherlekar, 2016).
Overall mutant and epistatic analyses presented in this study find a significant role for the F-BAR domain protein Synd in mediating pseudocleavage furrow extension. This study favors a model in which Synd, along with Anillin and RhoGEF2, provide a platform for recruitment of Dia and Pnut to allow persistent and stable growth of actin to promote furrow elongation. Further experiments combining protein interactions and deduction of the biophysical nature of Synd-Pnut-actin association with the plasma membrane will elucidate the molecular mechanism that makes Synd an important component of pseudocleavage furrow-extension or contractile-ring assembly at large (Sherlekar, 2016).
Membranes form elaborate structures that are highly tailored to their specialized cellular functions, yet the mechanisms by which these structures are shaped remain poorly understood. This study shows that the conserved membrane-remodeling C-terminal Eps15 Homology Domain (EHD) protein Past1 is required for the normal assembly of the subsynaptic muscle membrane reticulum (SSR) at the Drosophila melanogaster larval neuromuscular junction (NMJ). past1 mutants exhibit altered NMJ morphology, decreased synaptic transmission, reduced glutamate receptor levels, and a deficit in synaptic homeostasis. The membrane-remodeling proteins Amphiphysin and Syndapin colocalize with Past1 in distinct SSR subdomains, and collapse into Amphiphysin-dependent membrane nodules in the SSR of past1 mutants. These results suggest a mechanism by which the coordinated actions of multiple lipid-binding proteins lead to the elaboration of increasing layers of the SSR, and uncover new roles for an EHD protein at synapses (Koles, 2015).
Dozens of lipid-binding proteins dynamically remodel membranes, generating diverse cell shapes, sculpting organelles, and promoting traffic between subcellular compartments. Although the activities of many of these membrane-remodeling proteins have been studied individually, what is lacking is an understanding of how membrane-remodeling factors work together to generate specialized membranes in vivo (Koles, 2015).
C-terminal Eps15 Homology Domain (EHD)-family proteins encode large membrane-binding ATPases with structural similarity to dynamin and function at a variety of steps of membrane transport. These proteins contain an ATPase domain, a helical lipid-binding domain, and a carboxy-terminal EH domain that interacts with Asn-Pro-Phe (NPF)-containing binding partners. Although their mechanism of action is not fully understood, it is postulated that C-terminal EHD proteins bind and oligomerize in an ATP-dependent manner on membrane compartments, where they are involved in the trafficking of cargo. The mouse and human genomes each contain four highly similar EHD proteins (EHD1-4), which have both unique and overlapping functions. EHD proteins interact with several members of the Bin/Amphiphysin/Rvs167 (BAR) and Fes/Cip4 homology-BAR (F-BAR) protein families, which themselves can remodel membranes via their crescent-shaped dimeric BAR domains. In mammals, EHD proteins associate with the NPF motifs of the F-BAR proteins Syndapin I and II, and these interactions are critical for recycling of cargo from endosomes to the plasma membrane in cultured cells. In Caenorhabditis elegans, the sole EHD protein Rme-1 colocalizes and functions with the BAR protein Amphiphysin and the F-BAR protein Syndapin, also via their NPF motifs (Pant, 2009). Further, EHD1 has been suggested to drive the scission of endosomal recycling tubules generated by the membrane-deforming activities of Syndapin 2 and another NPF-containing protein, MICAL-L1. However, the combined membrane-remodeling activities that might arise in vivo from the shared functions of C-terminal EHD and NPF-containing proteins remain unclear (Koles, 2015).
The Drosophila neuromuscular junction (NMJ) is a powerful system in which to study membrane remodeling. On the postsynaptic side of the NMJ, a highly convoluted array of muscle membrane infoldings called the subsynaptic reticulum (SSR) incorporates neurotransmitter receptors, ion channels, and cell adhesion molecules. Assembly of the SSR during larval growth involves activity-dependent targeted exocytosis mediated by the small GTPase Ral and its effector, the exocyst complex, as well as the t-SNARE (target soluble N-ethylmaleimide-sensitive factor attachment protein receptor) receptor gtaxin/Syx18 and scaffolding proteins such as Discs Large (Dlg). Many proteins with predicted membrane-remodeling activities, including Drosophila homologues of Syndapin (Synd) and Amphiphysin (Amph), localize extensively to SSR membranes, making them prime candidates to facilitate SSR elaboration. Amph regulates the postsynaptic turnover of the trans-synaptic cell adhesion molecule FasII, but its role in organizing the SSR is unknown (Koles, 2015).
The Drosophila melanogaster genome encodes a single C-terminal EHD protein called Putative achaete/scute target (Past1). Past1 mutants exhibit defects in endocytic recycling in larval nephrocytes, sterility and aberrant development of the germline, and short lifespan, but the functions of Past1 at the NMJ have not been explored. Mammalian EHD1 localizes to the mouse NMJ, but its function there has been difficult to ascertain, perhaps due to redundancy with other EHD proteins. This study takes advantage of the fact that Past1 encodes the only Drosophila C-terminal EHD protein and define its role at the NMJ (Koles, 2015).
Putting together the current observations at the NMJ and in S2 cells with previous results from other groups, a new working model is proposed for how Past1 functions in synaptic membrane elaboration. The first key observation of this paper is the finding that Past1 is required for normal elaboration of the SSR and that this function depends on its ATP-binding and thus membrane-remodeling activity. Next it was found that in wild-type SSR, Amph localizes to a domain proximal to the bouton, whereas Past1 and Synd localize to a more extended tubulovesicular domain. By contrast, in the absence of Past1, the SSR rearranges into highly organized subdomains, with a core of Synd surrounded by a shell of Amph (likely corresponding to membrane sheets. Amph was found to be required for the formation of the sheets (perhaps by regulating the tight curvature at the tips of these membrane structures) and for consolidation of Synd into nodules. Further, FRAP data indicate that the nodules result in significantly increased membrane flow within the SSR relative to wild-type SSR, suggesting reduced complexity. Finally, S2 cell data indicate that Past1 may activate the membrane-binding/remodeling activity of Synd (Koles, 2015).
These results suggest a novel mechanism for SSR elaboration at the wild-type NMJ involving sequential steps of membrane remodeling. In this model, Amph localizes and generates membrane tubules proximal to the bouton, and Past1 and Synd work together to further elaborate the tubules distal to the bouton. Successive rounds of these events could lead to the growth and expansion of layers of reticulum. In past1 mutants, this process is severely compromised, resulting in nodules containing a core of inactive Synd packed by Amph-dependent membrane sheets (Koles, 2015).
One issue that remains to be resolved is whether direct physical interactions among Past1, Amph, and Synd (within the SSR subdomains to which they colocalize) contribute to Past1-dependent membrane remodeling at the NMJ, as they do in other systems. S2 cell data suggest that Past1 and Synd functionally interact in vivo. However, no Past1-Amph or Past1-Synd complexes were detected using coprecipitation experiments in extracts from Drosophila larvae or S2 cells or with purified proteins, suggesting that either they do not directly interact or their interactions are not preserved in solution under the conditions tested. Genetic experiments at the NMJ using mutations that disrupt putative Past1-Synd and Past1-Amph interactions are unlikely to be informative because synd and amph single mutants exhibit no dramatic phenotype in SSR organization, perhaps due to redundancy with other membrane-remodeling proteins. In fact, in addition to Amph and Synd, it was found that the BAR proteins Cip4 and dRich are localized to nodules in past1 mutants, suggesting that multiple membrane-remodeling proteins are available to function in the Past1-dependent pathway. In the future, it will be important to build into the working model the additional roles of these and other SSR-localized membrane-remodeling proteins, as well as the timing of exocyst-dependent membrane addition (Koles, 2015 and references therein).
The results demonstrate that postsynaptic Past1 plays critical roles in the structure and function of the Drosophila NMJ. Past1 mutant NMJs exhibit aberrant morphology and excess ghost boutons. These ghost boutons are unlikely to be due to defective clearance of excess neuronal membrane, since large amounts of neuronal debris were not observed. They are also unlikely to be related to excess ghost boutons seen in Wingless (Wg) signaling pathway mutants, since past1 mutants do not phenocopy many other aspects of reduced Wg signaling, including increased GluR levels, disrupted presynaptic function, and reduction in bouton number. The likeliest interpretation is that Past1 functions directly in SSR membrane elaboration, consistent with EM observations, and ghost boutons may arise when membrane nodules become too severe to allow SSR assembly around boutons that form toward the end of larval development (Koles, 2015).
Another prominent synaptic phenotype that found in past1 mutants is a strong and specific reduction in localization of GluRIIA to postsynaptic specializations, resulting in decreased mEPSP amplitude. This decrease in GluRIIA could potentially arise by many mechanisms, including altered transcriptional or translational regulation or GluR traffic to or from the synapse. Indeed, expression of a dominant-negative EHD1 suppresses AMPA glutamate receptor recycling in hippocampal dendritic spines. Although there has been little evidence that Drosophila GluRs are regulated by membrane traffic, the data implicating the membrane-remodeling protein Past1 indicate that this may be the case. Finally, unlike the great majority of perturbations that reduce GluRIIA levels, past1 mutants surprisingly fail to compensate for this loss by homeostatic up-regulation of presynaptic release, suggesting that Past1 could be involved in relaying an as-yet-unidentified retrograde signal for synaptic homeostasis. Further work exploring mechanisms of GluRIIA regulation and retrograde signaling will be required to understand the role of Past1 in these events (Koles, 2015).
The present data cannot distinguish whether the function of Past1 in GluR traffic or homeostasis is directly related to its role in SSR elaboration, and it is possible that membrane compartments independent of the SSR are required for these functions and are disrupted in the mutant. The finding that GluRIIA levels are still reduced in amph; past1 double mutants although SSR nodules are suppressed supports the conclusion that GluR localization defects are independent of aberrant SSR morphogenesis. Of note, many mutants with severely defective SSR and/or reduced GluR levels exhibit normal homeostasis (e.g., GluRIIA, which also has reduced SSR, Dlg, Gtaxin, and Pak1, suggesting that homeostasis is a specific function of Past1 rather than a general SSR- or GluR-related defect (Koles, 2015).
Past1 represents the sole EHD homologue in Drosophila, whereas mammals express four EHD proteins with distinct functions. Of importance, many of the roles for EHD proteins at the NMJ and in muscle are likely to be conserved. Past1 localizes to the NMJ, the muscle cortex, and myotendinous junctions. However, unlike EHD1, Past1 does not significantly localize to t-tubules. The activities identified for Past1 at the Drosophila NMJ may inform mechanisms by which EHD2 participates in sarcolemmal repair at the muscle cortex, EHD3 functions in cardiac muscle physiology, and EHD1 and EHD4 act at the mouse NMJ (Mate, 2012). The current findings set the stage for uncovering how neuromuscular synapses are formed and elaborated and illustrate how cooperation between lipid-remodeling proteins can create highly complex membrane structures (Koles, 2015).
Cytokinesis is a highly ordered cellular process driven by interactions between central spindle microtubules and the actomyosin contractile ring linked to the dynamic remodelling of the plasma membrane. The mechanisms responsible for reorganizing the plasma membrane at the cell equator and its coupling to the contractile ring in cytokinesis are poorly understood. This study reports that Syndapin, a protein containing an F-BAR domain required for membrane curvature, contributes to the remodelling of the plasma membrane around the contractile ring for cytokinesis. Syndapin colocalizes with phosphatidylinositol 4,5-bisphosphate (PI(4,5)P(2)) at the cleavage furrow, where it directly interacts with a contractile ring component, Anillin. Accordingly, Anillin is mislocalized during cytokinesis in Syndapin mutants. Elevated or diminished expression of Syndapin leads to cytokinesis defects with abnormal cortical dynamics. The minimal segment of Syndapin, which is able to localize to the cleavage furrow and induce cytokinesis defects, is the F-BAR domain and its immediate C-terminal sequences. Phosphorylation of this region prevents this functional interaction, resulting in reduced ability of Syndapin to bind to and deform membranes. Thus, the dephosphorylated form of Syndapin mediates both remodelling of the plasma membrane and its proper coupling to the cytokinetic machinery (Takeda, 2013).
This study provides the first compelling description of the requirements for an F-BAR protein in cytokinesis in animal cells. This work does not exclude other F-BAR proteins from participating in cytokinesis, but it does show a positive role for Syndapin in cortical membrane dynamics at the cleavage furrow. Syndapin's localization to the cleavage furrow and its in vitro membrane binding and tubulation are regulated by phosphorylation. The defects in cytokinesis ensuing from phosphomimetic mutants imply that phosphorylation of Syndapin regulates cytokinesis by affecting its membrane association. However, this does not exclude a possible indirect effect whereby phosphorylation may influence the association between Syndapin's SH3 and F-BAR domains, as has been proposed to auto-inhibit its membrane association. These findings would suggest that auto-inhibition results in reduced membrane binding, and yet no increased membrane binding and tubulation is seen with the full-length 12ST>A mutant. This implies that the major effect of phosphorylation is directly upon its membrane association. The phosphorylation of Syndapin could be a mechanism to prevent its premature association with the membrane at the cleavage furrow, as with phosphoregulation of Cdc15p during cytokinesis. Thus, Syndapin joins the F-BAR proteins of S. pombe and S. cerevisiae (Cdc15p and Hof1p, respectively) as proteins that are also phosphoregulated during cytokinesis (Takeda, 2013).
Syndapin's localization to the cleavage furrow requires its association with anionic lipids via its F-BAR domain. Syndapin also colocalizes with and directly binds to Anillin, but this interaction is dispensable for Syndapin localization. By contrast, Anillin is mislocalized during cytokinesis at least in primary spermatocytes of Syndapin mutants. Together these results led to a hypothesis that Syndapin may be a component of the coupling between the plasma membrane and the Anillin ring and hence the contractile ring. Alternatively, as Anillin itself has been proposed to have a role in linking the plasma membrane and the contractile ring, it is possible that Syndapin and Anillin share redundant function, and they may function cooperatively at the interface of plasma membrane and the contractile ring. Other candidate proteins for linking the contractile ring to the plasma membrane in cytokinesis are the C1 domain-containing MgcRacGAP of human cells and the C2 domain-containing protein Inn1 of budding yeast. Interestingly, Inn1 interacts with the F-BAR protein, Hof1p, and together they may cooperatively regulate membrane dynamics during cytokinesis in this organism. The Drosophila genome encodes several uncharacterized C2 domain-containing proteins, and it will be interesting to examine whether any of these proteins function cooperatively with Syndapin in cytokinesis. It is also possible that some of the other Drosophila F-BAR proteins (Cip4, Nwk, FCHo/CG8176, Fps85D and NOSTRIN/CG42388) function in cytokinesis. Such proteins could provide some functional redundancy to the molecular mechanism. Indeed, it cannot be excluded that other molecular components can participate in safeguarding the linkage of the membrane to the contractile ring. The importance of such molecules might vary between tissues, thus accounting for the differences in severity of Syndapin phenotypes between different cell types (Takeda, 2013).
Structure-function analyses demonstrated that expression of Syndapin fragments comprising the minimum segments required for the cleavage furrow localization (i.e. the F-BAR domain plus its C-terminal 65 amino acids) could induce dominant, strong cytokinesis defects. Expression of exogenous Syndapin also induced similar abnormal cortical behaviour with furrow ingression failure and severe generalized blebbing. Surprisingly, despite the robustness of these cytokinesis defects, the localization of the major components of the central spindle (Pavarotti) was not affected, although contractile ring components were misplaced around the central part of the cell. By contrast, expression of Syndapin segments that fail to bind to anionic lipids and localize to the cleavage furrow (i.e. ΔFBAR and K5E) did not affect cytokinesis. These results suggest that Syndapin affects cortical dynamics during cytokinesis by directly associating with anionic lipids on the plasma membrane. The S. pombe F-BAR protein, Cdc15p, has roles in organizing membrane domains into lipid rafts as well as in the contractile ring formation. Thus, it will be of future interest to determine whether Syndapin has an equivalent role in organizing membrane during cytokinesis in animal cells as a means of regulating cortical stiffness and dynamics (Takeda, 2013).
Syndapin is required for synaptic vesicle recycling both in mice and in flies, and an involvement in neuronal morphogenesis is regulated by developmentally controlled phosphorylation. This raises the question of whether the functions of Syndapin in synaptic vesicle trafficking and other developmental processes might follow similar regulatory processes. The shape of a membrane can be described by the radius of curvature in two perpendicular arcs. At the cleavage furrow, the radius of curvature along the axis of cell division will be positive, and perpendicular to this it will be negative. Similar curvatures will arise during vesicle recycling at the interface between the cap of a nascent vesicle and its parent membrane. The banana-shaped structure of F-BAR domains may make them ideal for associating with membrane in the context of such curvature, provided that all molecules orient in the one direction. An involvement of Syndapin in both cytokinesis and synaptic vesicle recycling would suggest that it can generate or stabilize varying degrees of positive curvature. When overexpressed in D.Mel-2 cells, narrow tubules decorated by Syndapin were sometime seen, and in vitro tubules were observed with approximate diameter of 55 nm. However, the diameter of positive curvature of a cleavage furrow will be at least one order of magnitude greater than this. Either Syndapin participates in forming smaller buds that become incorporated into the cleavage furrow or it indeed associates with membranes having larger diameters of curvature than may be suggested by the diameter of the concave face of its F-BAR domain. This latter possibility has some credibility because the extent of curvature will depend on the local membrane concentration of the F-BAR domain, and it would not be expected for the membrane to be saturated with the protein in vivo (otherwise, other membrane interacting proteins would be outcompeted). Thus, only narrow tubules are expected to be formed either in vitro or, as a result of overexpression, in vivo when membrane sites could be saturated (Takeda, 2013).
Several future challenges lay ahead beforethe regulation and roles of Syndapin in cytokinesis can be fully understood. Although the OA sensitivity of the protein phosphatase that dephosphorylates Syndapin suggests it is in the PP1 family, further studies are required to identify precisely the protein phosphatase(s) involved. Similarly, future studies will be necessary to identify the kinase(s) required for Syndapin's phosphoregulation. An understanding of Syndapin's precise cytokinetic role will be aided by more detailed description of its interacting partners. Although Syndapin interacts with Anillin, a full description of its functions in cytokinetic network is still needed. Only with this knowledge will it be understood how it might contribute to the coupling between the contractile ring and central spindle MTs underlying the cleavage furrow and the invaginating membrane (Takeda, 2013).
Syndapins belong to the F-BAR domain protein family whose predicted functions in membrane tubulation remain poorly studied in vivo. At Drosophila neuromuscular junctions, Syndapin is associated predominantly with a tubulolamellar postsynaptic membrane system known as the subsynaptic reticulum (SSR). syndapin overexpression greatly expands this postsynaptic membrane system. Syndapin can expand the SSR in the absence of dPAK and Dlg, two known regulators of SSR development. Syndapin's N-terminal F-BAR domain, required for membrane tubulation in cultured cells, is required for SSR expansion. Consistent with a model in which Syndapin acts directly on postsynaptic membrane, SSR expansion requires conserved residues essential for membrane binding in vitro. However, Syndapin's Src homology (SH) 3 domain, which negatively regulates membrane tubulation in cultured cells, is required for synaptic targeting and strong SSR induction. These observations advance knowledge of Syndapin protein function by 1) demonstrating the in vivo relevance of membrane remodeling mechanisms suggested by previous in vitro and structural analyses; 2) showing that SH3 domains are necessary for membrane expansion observed in vivo, and 3) confirming that F-BAR proteins control complex membrane structures (Kumar, 2009b).
Structural, cell biological, and in vitro studies of F-BAR domain proteins have contributed significantly toward a molecular understanding of how these proteins interact with membranes and other endocytic proteins to generate membrane tubules. However, knowledge of these proteins in vivo, in the multicellular context, remains very limited. This study provide an in vivo analyses of syndapin in the context of its role in the biogenesis of subsynaptic reticulum, a unusual complex membrane system. This study made three key observations on Drosophila syndapin: 1) it was shown that syndapin can promote formation of a tubulolamellar membrane system in vivo, 2) syndapin causes membrane remodeling that can occur without accompanying membrane fission, and 3) syndapin promotes SSR expansion by using evolutionarily conserved amino acids in its F-BAR domain. These three points are considered in turn below (Kumar, 2009b).
The SSR is a unique system of tubules and lamellae formed by extensive infoldings of the postsynaptic muscle membrane; thus, in organization, they are quite different from the relatively simple F-BAR-induced membrane tubules described in cultured cells. The SSR surrounds large boutons at the Drosophila NMJ. Although, a role for signaling and scaffolding proteins such as dPAK and Dlg has been demonstrated in the formation of SSR, mechanisms that underlie biogenesis of this complex membrane system are still poorly understood (Kumar, 2009b).
Syndapin overexpression in muscle caused induction of synaptic and extrasynaptic membrane-dense subsynaptic reticulum, based on optical and electron microscopic analyses. In particular, EM sections of membrane structures induced by syndapin overexpression showed not only circular/elliptical profiles expected for tubules, but also longer parallel membrane profiles, suggesting sections through lamellae as indeed is seen for native SSR. This is the first demonstration that an F-BAR protein can promote the formation of lamellar membrane infoldings. The mechanism by which syndapin may promote lamella formation is unclear, but it is likely that this arises from context specific interactions with other protein components of the SSR (Kumar, 2009b).
Syndapin seems to induce SSR through mechanisms that are either downstream of, or independent of, dPAK and Dlg function. This is indicated by three observations. First, the syndapin immunoreactivity is significantly reduced in dPAK and Dlg mutants. Second, unlike Dlg that can induce SSR when expressed either pre- or postsynaptically, syndapin acts in a cell-autonomous manner in postsynaptic muscle. This could indicate either a function downstream of these signaling molecules or an entirely independent mechanism. The third observation is that although Synd-induced membrane is strikingly similar to the endogenous SSR in general appearance; it has some notable differences from the endogenous SSR in structure and composition. Synd-induced SSR has more densely packed membranes (∼30% more membrane layers per micrometer) and also contains lower amounts of Dlg and dPAK than the endogenous SSR. Both of these differences could conceivably arise from limiting amounts of Dlg, dPAK or some other factor(s) required for the precise organization of the SSR; however, the current data do not address this issue unequivocally (Kumar, 2009b).
Does SSR biogenesis induced by syndapin reflect its true physiological function rather than an interesting but physiologically irrelevant activity of the protein? The SSR remains normal in synd loss-of-function mutants. Although this could suggest that Synd has no physiological function in SSR biogenesis, an alternative possibility is that other postsynaptic F-BAR proteins compensate for the absence of syndapin. Indeed, potential functional redundancies among F-BAR proteins are suggested by reports that different F-BAR proteins can coexist on a single tubule (Kumar, 2009b).
Although alternative models are tenable, it is suggested that Synd has a role in SSR biogenesis in vivo based on four arguments. First, Synd is localized to the postsynaptic SSR and would therefore most simply be expected to have an SSR-related function. The observed expansion of the SSR is consistent with this premise. Second, full-length Synd overexpression does not cause random patches of SSR to be induced all over the muscle surface but rather causes local expansions as well as flares that often seem to emanate from the existing SSR. Thus, the observed consequence of Synd overexpression seems to originate from sites to which Synd is normally targeted in vivo. Third, that Synd can promote formation of a unique, highly complex membrane system in vivo indicates that it participates in intricate processes that likely require the coordinated function of many different proteins. Finally, consistent with the previous argument, muscle expression of other Drosophila F-BAR domain proteins such as Nervous wreck does not induce SSR expansion (Kumar, 2009b).
In cultured cells, overexpression of F-BAR proteins induces transient, dynamin-containing plasma membrane tubules that are rapidly fragmented by dynamin-mediated membrane scission. Here, tubulation can be decoupled from membrane fission only if either the SH3 domain is removed or if SH3 interacting molecules (e.g., dynamin) are inhibited. A physiological decoupling of membrane tubulation and fission activities has been shown previously for the N-BAR domain proteins, mouse Amphiphysin 2 and Drosophila Amphiphysin, during T-tubule formation. The current observations suggest that similar physiological decoupling of the two activities—membrane deformation and membrane fission also occurs for the syndapin, an F-BAR domain protein (Kumar, 2009b).
In support of this, it was shown that unlike synd, the membrane fission protein dynamin is not enriched in the SSR; this is different from strong colocalization observed between dynamin and Syndapin in transient tubules in cultured cells. Furthermore, the presence of the dynamin-interacting SH3 domain does not inhibit Syndapin's ability to promote SSR formation. Thus, in contrast to prior observations in cultured cells, the data show that Syndapin in vivo can 1) be present without accompanying dynamin and 2) can form stable membrane infoldings without need to experimentally inhibit SH3 domain interactions (Kumar, 2009b).
The mechanism by which Syndapin promotes SSR formation is likely to require direct membrane interactions mediated by previously identified residues on the concave face of its F-BAR domain. Mutations in key residues on the concave face of the F-BAR domain, required for phospholipid binding, block the ability of Syndapin to induce SSR. Thus, mechanisms that underlie F-BAR protein's ability to tubulate membrane in vitro seem to be required for Syndapin's ability to expand the SSR. However, Synd-induced SSR formation requires additional events, including correct targeting to the postsynapse, a function that requires the C-terminal SH3 domain (Kumar, 2009b).
Although Synd lacking its SH3 domain is extremely efficient at membrane tubulation/remodeling in S2 cells, this truncated protein is not postsynaptically targeted in muscle cells and is ineffective for SSR expansion. Thus, the SH3 domain of Synd must interact with targeting molecules that control Syndapin's postsynaptic localization. By extension, the targeting of other F-BAR domain proteins, which may be mediated by analogous SH3 domain interactions, could be important for their respective in vivo functions (Kumar, 2009b).
Could these observations on Syndapin be relevant to the function of other F-BAR domain proteins? The SSR has some similarity to plasma membrane specializations such as the demarcation membrane system of megakaryocytes, which give rise to platelet plasma membrane. It is conceivable that other F-BAR domain proteins will be found to be involved in the biogenesis of these or other complex membrane system. Further studies are required to understand the different processes involved in SSR biogenesis and also to test the relevance of these findings to other members of the F-BAR protein family (Kumar, 2009b).
Syndapin is a conserved dynamin-binding protein, with predicted function in synaptic-vesicle endocytosis. This study combined genetic mutational analysis with in vivo cell biological assays to ask whether Drosophila Syndapin (Synd) is an essential component of synaptic-vesicle recycling. The only isoform of Drosophila Syndapin (Synd) is broadly expressed and at high levels in the nervous system. synd mutants are late-larval lethals, but fertile adult 'escapers' frequently emerge. Contrary to expectation, this study reports that the Synd protein is predominantly postsynaptic, undetectable at presynaptic varicosities at Drosophila third-instar larval neuromuscular junctions. Electrophysiological and synaptopHluorin imaging in control, synd-deficient or synd-overexpressing motor neurons reveals that synd is dispensable for synaptic-vesicle endocytosis. This work in Drosophila leads to the suggestion that Syndapin may not be a general or essential component in dynamin-dependent synaptic-vesicle endocytosis (Kumar, 2009a).
Syndapin/Pascin family F-BAR domain proteins bind directly to membrane lipids and are associated with actin dynamics at the plasma membrane. Previous reports have also implicated mammalian syndapin 2 in endosome function during receptor recycling, but precise analysis of a putative recycling function for syndapin in mammalian systems is difficult because of syndapin effects on the earlier step of endocytic uptake, and potential redundancy among the three separate genes that encode mammalian syndapin isoforms. This study analyzed the endocytic transport function of the only C. elegans syndapin, SDPN-1. SDPN-1 is a resident protein of the early and basolateral recycling endosomes in the C. elegans intestinal epithelium, and sdpn-1 deletion mutants display phenotypes indicating a block in basolateral recycling transport. sdpn-1 mutants accumulate abnormal endosomes positive for early endosome and recycling endosome markers that are normally separate, and such endosomes accumulate high levels of basolateral recycling cargo. Furthermore, strong colocalization of endosomal SDPN-1 was observed with the F-actin biosensor Lifeact; loss of SDPN-1 greatly reduced Lifeact accumulation on early endosomes. Taken together these results provide strong evidence for an in vivo function of syndapin in endocytic recycling, and suggest that syndapin promotes transport via endosomal fission (Gleason, 2016).
Insights into mechanisms coordinating membrane remodeling, local actin nucleation, and postsynaptic scaffolding during postsynapse formation are important for understanding vertebrate brain function. Gene knockout and RNAi in individual neurons reveal that the F-BAR protein syndapin I is a crucial postsynaptic coordinator in formation of excitatory synapses. Syndapin I deficiency caused significant reductions of synapse and dendritic spine densities. These syndapin I functions reflected direct, SH3 domain-mediated associations and functional interactions with ProSAP1/Shank2. They furthermore required F-BAR domain-mediated membrane binding. Ultra-high-resolution imaging of specifically membrane-associated, endogenous syndapin I at membranes of freeze-fractured neurons revealed that membrane-bound syndapin I preferentially occurred in spines and formed clusters at distinct postsynaptic membrane subareas. Postsynaptic syndapin I deficiency led to reduced frequencies of miniature excitatory postsynaptic currents, i.e., to defects in synaptic transmission phenocopying ProSAP1/Shank2 knockout, and impairments in proper synaptic ProSAP1/Shank2 distribution. Syndapin I-enriched membrane nanodomains thus seem to be important spatial cues and organizing platforms, shaping dendritic membrane areas into synaptic compartments (Schneider, 2014).
Glycine receptors (GlyRs) mediate inhibitory neurotransmission in spinal cord and brainstem. They are clustered at inhibitory postsynapses via a tight interaction of their beta subunits (GlyRbeta) with the scaffolding protein gephyrin. In an attempt to isolate additional proteins interacting with GlyRbeta, pulldown experiments were performed with rat brain extracts using a glutathione S-transferase fusion protein encompassing amino acids 378-455 of the large intracellular loop of GlyRbeta as bait. This identified syndapin I (SdpI) as a novel interaction partner of GlyRbeta that coimmunoprecipitates with native GlyRs from brainstem extracts. Both SdpI and SdpII bound efficiently to the intracellular loop of GlyRbeta in vitro and colocalized with GlyRbeta upon coexpression in COS-7 cells. The SdpI-binding site was mapped to a proline-rich sequence of 22 amino acids within the intracellular loop of GlyRbeta. Deletion and point mutation analysis disclosed that SdpI binding to GlyRbeta is Src homology 3 domain-dependent. In cultured rat spinal cord neurons, SdpI immunoreactivity was found to partially colocalize with marker proteins of inhibitory and excitatory synapses. When SdpI was acutely knocked down in cultured spinal cord neurons by viral miRNA expression, postsynaptic GlyR clusters were significantly reduced in both size and number. Similar changes in GlyR cluster properties were found in spinal cultures from SdpI-deficient mice. These results are consistent with a role of SdpI in the trafficking and/or cytoskeletal anchoring of synaptic GlyRs (Del Pino, 2014).
Syndapin 1 FBAR, a member of the Bin-amphiphysin-Rvs (BAR) domain protein family, is known to induce membrane curvature and is an essential component in biological processes like endocytosis and formation and growth of neurites. This study quantified the curvature sensing of FBAR on reconstituted porcine brain lipid vesicles and shows that it senses membrane curvature at low density whereas it induces and reinforces tube stiffness at higher density. FBAR strongly up-concentrates on the high curvature tubes pulled out of Giant Unilamellar lipid Vesicles (GUVs); this sorting behavior is strongly amplified at low protein densities. Interestingly, FBAR from syndapin 1 has a large affinity for tubular membranes with curvatures larger than its own intrinsic concave curvature. Finally, the effect of FBAR on membrane relaxation kinetics were studied with high temporal resolution; the protein was found to increase relaxation time of the tube holding force in a density-dependent fashion (Ramesh, 2013).
Spatial control of cortical actin nucleation is indispensable for proper establishment and plasticity of cell morphology. Cobl is a novel WH2 domain-based actin nucleator. The cellular coordination of Cobl's nucleation activity, however, has remained elusive. This study reveals that Cobl's cellular functions are dependent on syndapin. Cobl/syndapin complexes form in vivo, as demonstrated by colocalization, coimmunoprecipitation and subcellular recruitment studies. In vitro reconstitutions and subcellular fractionations demonstrate that, via its lipid-binding Fer/CIP4 Homology (FCH)-Bin/Amphiphysin/Rvs (F-BAR) domain, syndapin recruits Cobl to membranes. Consistently, syndapin I RNAi impairs cortical localization of Cobl. Further functional studies in neurons show that Cobl and syndapin I work together in dendritic arbor development. Importantly, both proteins are crucial for dendritogenesis. Cobl-mediated functions in neuromorphogenesis critically rely on syndapin I and interestingly also on Arp3. Endogenous Cobl, syndapin I and the Arp2/3 complex activator and syndapin-binding partner N-WASP were present in one complex, as demonstrated by coimmunoprecipitations. Together, these data provide detailed insights into the molecular basis for Cobl-mediated functions and reveal that different actin nucleators are functionally intertwined by syndapin I during neuromorphogenesis (Schwintzer, 2011).
Members of the Bin/amphiphysin/Rvs (BAR) domain protein superfamily are involved in membrane remodeling in various cellular pathways ranging from endocytic vesicle and T-tubule formation to cell migration and neuromorphogenesis. Membrane curvature induction and stabilization are encoded within the BAR or Fer-CIP4 homology-BAR (F-BAR) domains, alpha-helical coiled coils that dimerize into membrane-binding modules. BAR/F-BAR domain proteins often contain an SH3 domain, which recruits binding partners such as the oligomeric membrane-fissioning GTPase dynamin. How precisely BAR/F-BAR domain-mediated membrane deformation is regulated at the cellular level is unknown. This study presents the crystal structures of full-length syndapin 1 and its F-BAR domain. The data show that syndapin 1 F-BAR-mediated membrane deformation is subject to autoinhibition by its SH3 domain. Release from the clamped conformation is driven by association of syndapin 1 SH3 with the proline-rich domain of dynamin 1, thereby unlocking its potent membrane-bending activity. It is hypothesized that this mechanism might be commonly used to regulate BAR/F-BAR domain-induced membrane deformation and to potentially couple this process to dynamin-mediated fission. These data thus suggest a structure-based model for SH3-mediated regulation of BAR/F-BAR domain function (Rao, 2010).
Coordinated functions of the actin cytoskeleton and microtubules, which require careful control in time and space, are indispensable for the drastic alterations of neuronal morphology during neuromorphogenesis and neuronal network formation. Actin filament formation driven by the Arp2/3 complex and its activator neural Wiskott-Aldrich syndrome protein (N-WASP) is important for proper axon development. The underlying molecular mechanisms for targeting to and specific activation of N-WASP at the neuronal plasma membrane, however, have thus far remained elusive. This study shows that syndapin I is critical for proper neuromorphogenesis and uses N-WASP as a cytoskeletal effector. Upon N-WASP binding, syndapins release N-WASP autoinhibition. Syndapins cooperate with Cdc42 and phosphatidyl-inositol-(4,5)-bisphosphate. Syndapins furthermore specifically bind to phosphatidylserine-containing membranes via their extended F-BAR domain. Dissecting the syndapin functions actin nucleation and direct membrane binding in vivo, it was demonstrated that both functions are physiologically relevant and required. Constitutive plasma membrane-targeting experiments in vivo indicate that specifically actin nucleation at the cell cortex is triggered by syndapins. Consistent with syndapins steering N-WASP as downstream effector for cortical actin nucleation, syndapin-induced neuronal arborization is N-WASP and Cdc42 dependent. The functions of syndapin-N-WASP complexes in neuromorphogenesis were revealed by loss-of-function studies. Knockdown of syndapin I leads to impaired axon development and especially phenocopies the aberrant axon branching observed upon N-WASP and Arp2/3 complex deficiency. In contrast, proper length control involves another N-WASP-binding protein, Abp1. These data thus reveal that syndapin I is crucial for neuromorphogenesis and that different N-WASP activators ensure fine control of N-WASP activity and have distinct functions during neuronal network formation (Dharmalingam, 2009).
Syndapin I (SdpI) interacts with proteins involved in endocytosis and actin dynamics and was therefore proposed to be a molecular link between the machineries for synaptic vesicle recycling and cytoskeletal organization. This study reports the identification and characterization of SdpII, a ubiquitously expressed isoform of the brain-specific SdpI. Certain splice variants of rat SdpII in other species were named FAP52 and PACSIN 2. SdpII binds dynamin I, synaptojanin, synapsin I, and the neural Wiskott-Aldrich syndrome protein (N-WASP), a stimulator of Arp2/3 induced actin filament nucleation. In neuroendocrine cells, SdpII colocalizes with dynamin, consistent with a role for syndapin in dynamin-mediated endocytic processes. The src homology 3 (SH3) domain of SdpI and -II inhibited receptor-mediated internalization of transferrin, demonstrating syndapin involvement in endocytosis in vivo. Overexpression of full-length syndapins, but not the NH(2)-terminal part or the SH3 domains alone, had a strong effect on cortical actin organization and induced filopodia. This syndapin overexpression phenotype appears to be mediated by the Arp2/3 complex at the cell periphery because it was completely suppressed by coexpression of a cytosolic COOH-terminal fragment of N-WASP. Consistent with a role in actin dynamics, syndapins localized to sites of high actin turnover, such as filopodia tips and lamellipodia. These results strongly suggest that syndapins link endocytosis and actin dynamics (Qualmann, 2000).
The GTPase dynamin has been clearly implicated in clathrin-mediated endocytosis of synaptic vesicle membranes at the presynaptic nerve terminal. This study describes a novel 52-kDa protein in rat brain that binds the proline-rich C terminus of dynamin. Syndapin I (synaptic, dynamin-associated protein I) is highly enriched in brain where it exists in a high molecular weight complex. Syndapin I can be involved in multiple protein-protein interactions via a src homology 3 (SH3) domain at the C terminus and two predicted coiled-coil stretches. Coprecipitation studies and blot overlay analyses revealed that syndapin I binds the brain-specific proteins dynamin I, synaptojanin, and synapsin I via an SH3 domain-specific interaction. Coimmunoprecipitation of dynamin I with antibodies recognizing syndapin I and colocalization of syndapin I with dynamin I at vesicular structures in primary neurons indicate that syndapin I associates with dynamin I in vivo and may play a role in synaptic vesicle endocytosis. Furthermore, syndapin I associates with the neural Wiskott-Aldrich syndrome protein, an actin-depolymerizing protein that regulates cytoskeletal rearrangement. These characteristics of syndapin I suggest a molecular link between cytoskeletal dynamics and synaptic vesicle recycling in the nerve terminal (Qualmann, 1999).
Search PubMed for articles about Drosophila Syndapin
Ahuja, R., Pinyol, R., Reichenbach, N., Custer, L., Klingensmith, J., Kessels, M. M. and Qualmann, B. (2007). Cordon-bleu is an actin nucleation factor and controls neuronal morphology. Cell 131(2): 337-350. PubMed ID: 17956734
de Kreuk, B. J., Nethe, M., Fernandez-Borja, M., Anthony, E. C., Hensbergen, P. J., Deelder, A. M., Plomann, M. and Hordijk, P. L. (2011). The F-BAR domain protein PACSIN2 associates with Rac1 and regulates cell spreading and migration. J Cell Sci 124(Pt 14): 2375-2388. PubMed ID: 21693584
Del Pino, I., Koch, D., Schemm, R., Qualmann, B., Betz, H. and Paarmann, I. (2014). Proteomic analysis of glycine receptor beta subunit (GlyRbeta)-interacting proteins: evidence for syndapin I regulating synaptic glycine receptors. J Biol Chem 289(16): 11396-11409. PubMed ID: 24509844
Dharmalingam, E., Haeckel, A., Pinyol, R., Schwintzer, L., Koch, D., Kessels, M. M. and Qualmann, B. (2009). F-BAR proteins of the syndapin family shape the plasma membrane and are crucial for neuromorphogenesis. J Neurosci 29(42): 13315-13327. PubMed ID: 19846719
Frost, A., Perera, R., Roux, A., Spasov, K., Destaing, O., Egelman, E. H., De Camilli, P. and Unger, V. M. (2008). Structural basis of membrane invagination by F-BAR domains. Cell 132(5): 807-817. PubMed ID: 18329367
Gleason, A. M., Nguyen, K. C., Hall, D. H. and Grant, B. D. (2016). Syndapin/SDPN-1 is required for endocytic recycling and endosomal actin association in the C. elegans intestine. Mol Biol Cell. PubMed ID: 27630264
Koles, K., Messelaar, E. M., Feiger, Z., Yu, C. J., Frank, C. A. and Rodal, A. A. (2015). The EHD protein Past1 controls postsynaptic membrane elaboration and synaptic function. Mol Biol Cell 26(18):3275-88. PubMed ID: 26202464
Kostan, J., Salzer, U., Orlova, A., Toro, I., Hodnik, V., Senju, Y., Zou, J., Schreiner, C., Steiner, J., Merilainen, J., Nikki, M., Virtanen, I., Carugo, O., Rappsilber, J., Lappalainen, P., Lehto, V. P., Anderluh, G., Egelman, E. H. and Djinovic-Carugo, K. (2014). Direct interaction of actin filaments with F-BAR protein pacsin2. EMBO Rep 15(11): 1154-1162. PubMed ID: 25216944
Kumar, V., Alla, S. R., Krishnan, K. S. and Ramaswami, M. (2009a). Syndapin is dispensable for synaptic vesicle endocytosis at the Drosophila larval neuromuscular junction. Mol Cell Neurosci 40(2): 234-241. PubMed ID: 19059483
Kumar, V., Fricke, R., Bhar, D., Reddy-Alla, S., Krishnan, K. S., Bogdan, S. and Ramaswami, M. (2009b). Syndapin promotes formation of a postsynaptic membrane system in Drosophila. Mol Biol Cell 20(8): 2254-2264. PubMed ID: 19244343
Qualmann, B., Roos, J., DiGregorio, P. J. and Kelly, R. B. (1999). Syndapin I, a synaptic dynamin-binding protein that associates with the neural Wiskott-Aldrich syndrome protein. Mol Biol Cell 10(2): 501-513. PubMed ID: 9950691
Qualmann, B. and Kelly, R. B. (2000). Syndapin isoforms participate in receptor-mediated endocytosis and actin organization. J Cell Biol 148(5): 1047-1062. PubMed ID: 10704453
Ramachandran, P., Barria, R., Ashley, J. and Budnik, V. (2009). A critical step for postsynaptic F-actin organization: regulation of Baz/Par-3 localization by aPKC and PTEN. Dev Neurobiol 69(9): 583-602. PubMed ID: 19472188
Ramesh, P., Baroji, Y. F., Reihani, S. N., Stamou, D., Oddershede, L. B. and Bendix, P. M. (2013). FBAR syndapin 1 recognizes and stabilizes highly curved tubular membranes in a concentration dependent manner. Sci Rep 3: 1565. PubMed ID: 23535634
Rao, Y., Ma, Q., Vahedi-Faridi, A., Sundborger, A., Pechstein, A., Puchkov, D., Luo, L., Shupliakov, O., Saenger, W. and Haucke, V. (2010). Molecular basis for SH3 domain regulation of F-BAR-mediated membrane deformation. Proc Natl Acad Sci U S A 107(18): 8213-8218. PubMed ID: 20404169
Roos, J. and Kelly, R. B. (1998). Dap160, a neural-specific Eps15 homology and multiple SH3 domain-containing protein that interacts with Drosophila dynamin. J Biol Chem 273(30): 19108-19119. PubMed ID: 9668096
Schneider, K., Seemann, E., Liebmann, L., Ahuja, R., Koch, D., Westermann, M., Hubner, C. A., Kessels, M. M. and Qualmann, B. (2014). ProSAP1 and membrane nanodomain-associated syndapin I promote postsynapse formation and function. J Cell Biol 205(2): 197-215. PubMed ID: 24751538
Schwintzer, L., Koch, N., Ahuja, R., Grimm, J., Kessels, M. M. and Qualmann, B. (2011). The functions of the actin nucleator Cobl in cellular morphogenesis critically depend on syndapin I. EMBO J 30(15): 3147-3159. PubMed ID: 21725280
Sherlekar, A. and Rikhy, R. (2016). Syndapin promotes pseudocleavage furrow formation by actin organization in the syncytial Drosophila embryo. Mol Biol Cell 27(13): 2064-2079. PubMed ID: 27146115
Simpson, F., Hussain, N. K., Qualmann, B., Kelly, R. B., Kay, B. K., McPherson, P. S. and Schmid, S. L. (1999). SH3-domain-containing proteins function at distinct steps in clathrin-coated vesicle formation. Nat Cell Biol 1(2): 119-124. PubMed ID: 10559884
Takeda, T., Robinson, I. M., Savoian, M. M., Griffiths, J. R., Whetton, A. D., McMahon, H. T. and Glover, D. M. (2013). Drosophila F-BAR protein Syndapin contributes to coupling the plasma membrane and contractile ring in cytokinesis. Open Biol 3(8): 130081. PubMed ID: 23926047
date revised: 30 January, 2018
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