the tiny protrusions emerging from dendrites known as dendritic spines are the primary subcellular locations of excitatory synapses in the mammalian central nervous system . dendritic spines are typically ~1 - 2  m in length and 0.51  m in width of the spine head , with diverse morphologies , such as mushroomlike , stubby , and thin spines . these structures are mainly supported by the f - actin cytoskeleton . thus , f - actin cytoskeletal proteins and regulators are important factors for generating dendritic spines . many membrane proteins and adaptor and signaling molecules are also involved in controlling dendritic spine formation and maintenance . the most popular mechanism is that dendritic filopodia serve as precursors for dendritic spine formation . interestingly , filopodia are ubiquitously found on various cell types . in contrast , dendritic spines are neuron - specific structures . thus , the transition from filopodia to spines should be controlled by neuron - specific factors . these proteins or responses directly or indirectly regulate f - actin rearrangement and dynamics to promote dendritic spine formation . studies of cytoskeleton - associated cortactin - binding protein 2 ( cttnbp2 ) and heparan sulfate transmembrane proteoglycan ( hspg ) syndecan-2 serve as examples for these two categories , respectively . cttnbp2 is a neuron - specific cytoskeleton - associated protein and that is enriched at dendritic spines of mature neurons . although syndecan-2 is widely expressed in many cell types , it is highly concentrated at synapses in neurons . syndecan-2 cooperates with other proteins to trigger neurotransmission through a neuron - specific signal to induce dendritic spine formation . genomic analyses of patients with autism spectrum disorders ( asds ) indicated that both cttnbp2 and syndecan-2 were associated with asds [ 4 , 5 ] . additionally , neurofibromin , cask , and vcp coordinate with syndecan-2 to control dendritic spinogenesis and were also associated with neurological disorders . these findings suggest that these genes are critical for neuronal function , likely through their regulation of dendritic spine formation . in this review , we will summarize the functions of these proteins in dendritic spinogenesis and use these proteins as examples to discuss how neuron - specific molecules coordinate with ubiquitously expressed proteins to generate neuron - specific signals for dendritic spine formation . syndecan-2 is a type i membrane protein with a heparan sulfate modification at its ectodomain ( figure 1 ) . in mammals , the syndecan protein family contains four members , syndecan-1 , syndecan-2 , syndecan-3 , and syndecan-4 . in rodent brains , syndecan-2 and syndecan-3 are the two major syndecans expressed in neurons with differential distribution ; syndecan-2 is highly concentrated at synapses , while syndecan-3 is distributed along the axonal shaft . syndecan-2 is involved in cell - cell and cell - matrix interactions through its heparan sulfate modification . it can also bind growth factors , such as fibroblast growth factor ( fgf ) and epidermal cell growth factor , and it acts as a coreceptor for these growth factors . syndecan-2 is broadly and dynamically expressed in several tissues and cell types [ 7 , 8 ] . during neural development , its expression gradually increases concurrent with synapse formation [ 8 , 9 ] . in mature neurons , such as cultured rat hippocampal neurons at 18 days after plating in vitro ( div ) or later more importantly , overexpression of syndecan-2 in immature rat hippocampal cultured neurons , such as 1 - 2 div , when endogenous syndecan-2 is not yet expressed , dendritic filopodia are massively induced at 4 - 5 div and dendritic filopodia are then transformed to dendritic spines at 8 - 9 div [ 9 , 11 ] . those dendritic spines are expected to be functional , as they are adjacent to the presynaptic marker synaptophysin based on confocal microscopy [ 11 , 12 ] . syndecan-2-induced dendritic spinogenesis serves as a model to explore the mechanisms underlying the initiation of dendritic spinogenesis ( namely , dendritic filopodia formation ) , the transition from filopodia to spines , and dendritic spine maturation and maintenance . the ectodomain of syndecan-2 heparan sulfate modification is involved in cell - cell and cell - matrix interactions . its transmembrane domain is required for homodimerization or oligomerization , which is critical for the protein - protein interactions of syndecan-2 . the cytoplasmic domain of syndecan-2 contains only 32 amino acid residues ( figure 1 ) . although it is short , it is divided into three motifs , conserved domain 1 ( c1 ) , the variable region ( v ) , and conserved domain 2 ( c2 ) . the c1 and c2 motifs are conserved among different syndecans , while the sequences of the v regions vary ( figure 1 ) . the c1 motif is essential for syndecan-2-induced dendritic filopodia formation of rat hippocampal cultured neurons , as the syndecan-2c1 mutant completely loses the ability to promote filopodia formation and spine formation at 5 as well as 9 div [ 11 , 15 ] . the c2 is required for the dendritic filopodia - spines transition and dendritic spine maintenance [ 15 , 16 ] . expression of the c2 deletion mutant syndecan-2c2 at 2 div promotes dendritic filopodia formation at 5 div . however , those filopodia are unable to transform into dendritic spine at 9 div [ 11 , 15 , 16 ] . these analyses indicate that the function of syndecan-2 in dendritic spinogenesis can be separated into two sequential steps , namely , filopodia and spine formation , which are controlled by two distinct motifs in syndecan-2 . because both c1 and c2 motifs are short and lack recognizable enzymatic domains , syndecan-2 binding partners have been identified to determine its molecular mechanism underlying dendritic spine formation . several direct binding partners ( summarized in table 1 ) have been identified for the c1 domains of syndecan-2 , including neurofibromin   and ezrin . , the interactions between syndecan-2 and neurofibromin and cask have been shown to be relevant in dendritic spine formation . because the cytoplasmic tail of syndecan-2 is very short , it is unlikely that a single syndecan-2 molecule can simultaneously interact with all of its binding partners . because the c1 and c2 motifs are involved in two sequential processes , it is likely that neurofibromin and cask sequentially interact with syndecan-2 . alternatively , it is possible that because syndecan-2 forms at least a dimer through its transmembrane domain , different syndecan-2 molecules in dimers or oligomers separately interact with neurofibromin and cask . this would suggest that syndecan-2 , neurofibromin , and cask form a single large complex . neurofibromin encoded by the neurofibromatosis type i ( nf1 ) gene is characterized by its rasgap- ( ras gtpase activating protein- ) related domain ( grd ) ( figure 2(a ) ) [ 2124 ] . similar to syndecan-2 , neurofibromin is widely expressed in different cell types , though its expression level is much higher in the nervous system . nf1 is one of the most common human inherited disorders featured by changes in skin pigmentation , benign tumor growth , and learning difficulty [ 26 , 27 ] . in addition to its ras activity , neurofibromin can increase camp concentration by activating adenylate cyclase . although the molecular mechanisms are less clear , the grd and c - terminal region of neurofibromin are required for camp pathway activation ( figure 2(a ) ) . both gs - dependent and gs - independent pathways are involved in neurofibromin - regulated adenylate cyclase activation . the camp pathway has been shown to be involved in learning and memory in drosophila   and dendritic spine formation in the mammalian nervous system . in a yeast two - hybrid screen using different fragments of neurofibromin as baits , syndecan-2 was identified as a neurofibromin binding partner . one is the jn fragment corresponding to amino acid residues 13571473 in the grd of human neurofibromin ; the other is the pn fragment containing amino acid residues 26192719 ( figure 2(a ) ) . in addition to biochemical studies demonstrating the direct interaction between syndecan-2 and neurofibromin , fluorescence immunostaining further demonstrated the colocalization of syndecan-2 and neurofibromin at synapses in cultured hippocampal neurons . moreover , both nf1 knockdown and haploinsufficiency reduce the density of dendritic spines in both rat hippocampal and mouse cortical cultured neurons and in brains [ 11 , 32 ] , consistent with a function of neurofibromin in regulating dendritic spine formation . one study examined syndecan-2 downstream signaling for triggering filopodia formation . using a panel of inhibitors to suppress various kinase activities , protein kinase a ( pka ) was identified to be required for syndecan-2-induced filopodia formation . combined with the analysis using different motif deletion mutants of syndecan-2 , we found that the c1 motif of syndecan-2 is essential for pka - dependent filopodia formation . because neurofibromin interacts with the c1 motif and also activates the camp pathway , cultured hippocampal neurons were then used to investigate whether neurofibromin mediates syndecan-2-induced filopodia formation . both nf1 knockdown and jn fragment expression , which acts as a dominant - negative to disrupt the interaction between endogenous neurofibromin and syndecan-2 , thus , neurofibromin mediates the signal from syndecan-2 to the camp pathway to initiate dendritic spinogenesis . because filopodia are supported by f - actin bundles , the syndecan-2-neurofibromin - camp pathway has to induce f - actin polymerization and bundle formation to promote dendritic filopodia formation . the ena ( enabled)/vasp ( vasodilator - stimulated phosphoprotein ) protein family is a group of f - actin regulators that initiate actin polymerization and bundling . pka phosphorylation promotes ena / vasp protein activity to regulate the f - actin cytoskeleton . upon syndecan-2 overexpression , ena / vasp phosphorylation increases , consistent with camp pathway activation moreover , disruption of ena / vasp activity impairs syndecan-2-induced dendritic filopodia formation . in summary , these studies indicate that syndecan-2 overexpression enhances the ability of neurofibromin to activate the pka pathway , which then induces the ena / vasp activity to promote f - actin bundling and filopodia formation . although the pka pathway is required for dendritic filopodia formation , increased intracellular camp concentrations alone can not induce dendritic filopodia formation , suggesting that other factor(s ) are involved . from an immunoprecipitation - mass spectrometry study , valosin - containing protein ( vcp , also known as p97 ) was identified as a neurofibromin - binding protein . the entire d1 and d2 atpase domains of vcp are required for the interaction with the leucine - rich domain ( lrd ) of neurofibromin . vcp is a causative gene of inclusion body myopathy associated with paget 's disease of bone and frontotemporal dementia ( ibmpfd ) . in addition , vcp mutations are associated with asds and amyotrophic lateral sclerosis [ 36 , 37 ] . a combination of human genetic studies , mouse genetic models , and cultured hippocampal and cortical neurons have indicated that neurofibromin interacts with vcp and guides vcp to promote dendritic spinogenesis . the roles of vcp and neurofibromin in dendritic spine formation may account for the neural phenotypes in patients with mutations in the nf1 and vcp genes . however , it is still unclear how vcp regulates dendritic spine formation . to fully address the molecular regulation of neurofibromin and vcp in dendritic spinogenesis , the function of the syndecan-2-neurofibromin interaction in dendritic spine formation is summarized in figure 2(b ) . mutations in the human cask gene result in x - linked mental retardation and microcephaly with pontine and cerebellar hypoplasia [ 3943 ] . cask belongs to the membrane - associated guanylate kinase ( maguk ) family and functions as a scaffold protein to interact with more than two dozen cellular proteins . it is widely distributed in neurons , including synapses , dendrites , axons , and soma . at synapses , pontine explants and rat hippocampal cultured neurons , cask knockdown impairs synapse formation at the pre- and postsynapse , respectively [ 16 , 45 ] . at presynaptic sites , it binds the membrane protein neurexin and other scaffold proteins , such as mint1 , mlin7 , and liprin , to control presynaptic button formation [ 4548 ] . cask uses its pdz domain at the postsynaptic site to interact with the c2 motif of syndecan-2 . in cultured hippocampal neurons , expression of the pdz alone of cask or the c - terminal tail of syndecan-2 that disrupts the interaction between endogenous cask and syndecan-2 reduces dendritic spine density , narrows spine heads , and shortens spine length at 18 div , suggesting that the cask - syndecan-2 interaction is critical for dendritic spine formation . to investigate whether cask is involved in dendritic spinogenesis initiation or dendritic spine stabilization , a time course study using a knockdown approach in cultured hippocampal neurons the time window of 1518 div covering the initiation toward maturation of dendritic spinogenesis was used for analysis . at 15 div , wild - type dendritic spines are immature , long , and thin , and they are present at a low density . as they mature at 18 div , dendritic spine density increases , spine length decreases , and spine width increases . compared to control neurons , cask knockdown does not affect spine density , length , or size at 15 div , suggesting that cask is not critical for dendritic spinogenesis initiation . at 18 div , the data indicate that cask is important for dendritic spine maturation , likely by linking the membrane protein syndecan-2 to the f - actin cytoskeleton via protein 4.1 to stabilize dendritic spines ( figure 3 ) . in human embryonic kidney hek293 cells , syndecan-2 however , these filopodia can not mature into spines . because neurofibromin and cask are also expressed in hek293 cells , the aforementioned studies can not explain why syndecan-2-induced dendritic spines are only present in neurons . a neuron - specific factor must be present to control dendritic spine formation . because neurotransmission is a neuron - specific event and because dendritic filopodia are able to receive neurotransmission signals from presynaptic buttons , neurotransmission seems a likely factor that triggers the filopodia - spines transformation in a neuron - specific manner . indeed , egta treatment to chelate extracellular calcium or ap5 treatment to block nmdar activity , a major neurotransmitter gated calcium channel , impairs the endogenous filopodia - spines transition at 1517 div and syndecan-2-induced filopodia - spines transition at 59 div . in syndecan-2-overexpressing neurons , intracellular calcium concentration is increased compared to control neurons at 5 div . this increase is due to nmdar - regulated calcium influx because ap5 treatment effectively reduced the intracellular calcium concentration induced by syndecan-2 . the c2 motif of syndecan-2 is required for syndecan-2 overexpression - induced calcium influx , suggesting that the interaction with cask is involved in calcium influx . previous studies have shown that cask interacts with mlin7 via the l27 domains in both proteins [ 5052 ] and that mlin7 interacts with the c - terminal tail of nmdar subunit 2b ( nmdar2b ) through its pdz domain . the interaction between syndecan-2 , cask , mlin7 , and nmdar2b facilitates nmdar localization to the tips of dendritic filopodia , where nmdar may be activated by presynaptic stimulation , namely , glutamate , and induce calcium influx . disruption of the syndecan-2 , cask , mlin7 , and nmdar complex by overexpressing the interacting domains impairs nmdar filopodial distribution , calcium influx , and the filopodia - spines transition , suggesting that syndecan-2 triggers calcium influx via the cask - mlin7-nmdar complex and induces the filopodia - spines transition ( figure 3 ) . the morphological feature of the filopodia - spines transition is dendritic spine head enlargement and spine length shortening . calcium is known to regulate f - actin dynamics in dendritic spines [ 5456 ] , and gelsolin is a calcium - activated f - actin regulator . gelsolin deficiency impairs filopodial retraction of developing neurons   and inhibits activity - dependent f - actin remodeling in mature dendritic spines . it is also critical for the filopodia - spines transition induced by the syndecan-2-cask - mlin7-nmdar complex , as gelsolin knockdown maintains syndecan-2-induced protrusions at the filopodial stage . it is possible that other calcium regulated f - actin regulators also act downstream of syndecan-2 to control the filopodia - spines transition . more investigations are required to further elucidate the regulation . through its interactions with intracellular binding partners , the ubiquitously expressing protein syndecan-2 modulates the f - actin cytoskeleton , triggers neurotransmission , and promotes neuron - specific synapse formation . from dendritic filopodia formation , filopodia - spines transition to dendritic spine maturation , syndecan-2 interacts with different binding partners to control f - actin behaviors . syndecan-2 first activates the pka pathway via neurofibromin to promote f - actin polymerization and bundling for dendritic filopodia formation . it recruits nmdar to filopodial tips through its interaction with the cask - mlin7 complex and increases the postsynaptic responsiveness to presynaptic stimulation . calcium influx induces f - actin cytoskeleton rearrangement to allow for the morphological change from filopodia to spines . to further promote dendritic spine maturation and maintenance , syndecan-2 binds to the protein 4.1 through interactions with cask . throughout the entire process , neuron specificity falls within nmdar - mediated calcium influx , which induces f - actin cytoskeleton remodeling to result in morphological changes to the dendritic spine . these studies provide a comprehensive example of how a neuron - specific ion channel coordinates with other adhesion molecules and synaptic proteins to control dendritic spine formation . to identify a neuron - specific f - actin regulator involved in dendritic spinogenesis , we searched the database and literature and focused on cortactin - binding protein 2 ( cttnbp2 ) . cttnbp2 gene encodes a brain - specific protein that interacts with the sh3 domain of cortactin through its proline - rich domain . cortactin promotes and stabilizes f - actin branching [ 63 , 64 ] and thus plays a critical role for dendritic spine morphological maintenance . because cortactin is a ubiquitously expressed protein , its function in controlling dendritic spinogenesis must be regulated by a neuron - specific factor . the specific expression of cttnbp2 in the brain makes it a good candidate to control cortactin in dendritic spinogenesis . furthermore , de novo mutations in the cttnbp2 gene have been repeatedly identified in asd patients [ 5 , 37 , 66 ] . in a genomic analysis covering 3871 asd patients , results indicated that cttnbp2 is a high - confidence risk factor for asds with a false discovery rate less than 0.05% . these genetic data support a critical role for cttnbp2 in brain development and function . in the expression tag sequence ( est ) database ( http://www.ncbi.nlm.nih.gov/ ) , three variants have been identified as cttnbp2 transcripts , namely , cttnbp2-short ( cttnbp2-s ) , cttnbp2-intron ( cttnbp2-i ) , and cttnbp2-long ( cttnbp2-l ) . based on the nucleotide sequence , the first 625 predicted amino acid residues are shared among all variants . using an antibody against the common region of the cttnbp variants , immunoblotting revealed that the short form of cttnbp2 is the predominant protein product in brains . it is still unclear whether the cttnbp2-i and cttnbp2-l variants play any role in neurons . therefore , mutation analysis of asd patients is meaningful when the mutation was located within the cttnbp2-s variant sequences . seven de novo asd mutations in the cttnbp2 gene have been identified in the exons encoding cttnbp2-s . to further explore the association of cttnbp2 with asd , these mutations should be investigated in detail to determine their effects on cttnbp2 molecular function and neuronal morphogenesis . analysis of the amino acid sequence of cttnbp2-s predicts a coiled - coil domain at the n - terminal region and proline - rich domain at the c - terminus . the n - terminal coiled - coil domain mediates cttnbp2-s homooligomerization and heterooligomerization of cttnbp2-s and the striatin family [ 68 , 69 ] . both endogenous cttnbp2-s and overexpressed myc - tagged cttnbp2-s were found to be highly concentrated at dendritic spines in mature cultured hippocampal neurons . immunofluorescence analysis of adult brains also indicated that cttnbp2-s colocalized with f - actin puncta in vivo , presumably to dendritic spines . cttnbp2-s is critical for dendritic spine formation , as cttnbp2 knockdown right before dendritic spinogenesis at 12 div reduces spine density and spine head width measured at 18 div . consistent with the morphological changes , the frequency of mepsc ( miniature excitatory postsynaptic synaptic current ) is lower in cttnbp2 knockdown neurons at 18 div . in addition to dendritic spine formation , cttnbp2-s is involved in dendritic spine maintenance , as cttnbp2-s knockdown in mature neurons , such as 20 div , still reduces dendritic spine density at 26 div . cortactin is required for cttnbp2-s 's regulation of dendritic spinogenesis , as a cttnbp2-s mutant that can not interact with cortactin can not rescue cttnbp2 knockdown - induced spine deficiency . moreover , fluorescence recovery after photobleaching ( frap ) analysis indicates that cttnbp2-s regulates cortactin mobility in mature dendritic spines . in the presence of cttnbp2-s , the data suggest that cttnbp2-s retains cortactin in dendritic spines and controls dendritic spine formation and maintenance . the striatin protein family contains three mammalian members , namely , striatin , zinedin , and sg2na . they function as b - type regulatory subunits of protein phosphatase 2a ( pp2a ) to control pp2a subcellular location and substrate specificity [ 70 , 71 ] . striatin protein distribution to synapses is mediated by its interactions with cttnbp2-s through the n - terminal coiled - coil domains of both cttnbp2-s and striatin family members . , cttnbp2-s regulates f - actin dynamics and pp2a signaling at dendritic spines . in cos cells , exogenous cttnbp2-s was unexpectedly associated with the microtubule cytoskeleton in addition to the cortactin - f - actin cytoskeleton . cell - matrix interactions influence the cytoskeleton association of cttnbp2-s . in cos cells , cttnbp2-s cttnbp2-s gradually shifts its preference to the microtubule cytoskeleton when establishing cell - matrix interactions . cttnbp2-s is highly concentrated at dendritic spines in mature neurons where cttnbp2-s interacts with f - actin cytoskeletons . in the premature stages when dendritic spines have not yet formed , cttnbp2-s is already expressed and forms puncta attached on microtubule bundles along the dendritic shaft . the mid domain is required for the association of microtubule , and the n - terminal coiled - coil domain is involved in cttnbp2-s oligomerization . oligomerization allows the cttnbp2-s oligomer to contain multiple microtubule binding sites to induce microtubule bundling . during the dendritic extension stage , cttnbp2-s knockdown or disruption of microtubule bundling by overexpression of the n - terminal coiled - coil domain or mid domain impairs dendritic arborization . the studies suggest that , in addition to controlling the f - actin cytoskeleton , cttnbp2-s regulates microtubule stability to influence dendrite morphology . the dual roles of cttnbp2-s in controlling f - actin and the microtubule cytoskeletons require further investigation . as a neuron - specific morphology regulator and a high - confidence risk factor for asds for instance , what is the molecular mechanism regulating the association between cttnbp2-s and f - actin and microtubules ? are the associations of cttnbp2-s with f - actin and microtubules mutually exclusive ? alternatively , can cttnbp2-s act as a bridge to link f - actin and microtubules ? only cultured hippocampal neurons have been examined in functional studies of cttnbp2-s . in the future , in vivo studies should be considered . particularly , to address the association of cttnbp2 with asds , a mouse genetic model is required . the impact of cttnbp2 asd mutations on the molecular function of cttnbp2-s , brain development , and cognition must be studied to further understand the biological significance of cttnbp2 . although hundreds of genes are involved in dendritic spine formation , they should be either neuron - specific or directly or indirectly controlled by or linked to neuron - specific signaling or proteins to specifically regulate dendritic spine formation in neurons . in this review , syndecan-2-induced dendritic spine formation and the role of cttnbp2-s in controlling neuronal morphology provide two distinct examples of how neuronal morphology can be regulated in a neuron - specific manner . understanding these regulations is crucial for basic research and for understanding neurological disorder etiology , which could contribute to potential therapeutic treatments of the diseases .