Adapter protein that may provide indirect link between the endocytic membrane traffic and the actin assembly machinery. May regulate the formation of clathrin-coated vesicles. Involved in endocytosis of integrin beta-1 (ITGB1) and transferrin receptor (TFR); internalization of ITGB1 as DAB2-dependent cargo but not TFR may involve association with DAB2.
Endocytic adaptor proteins facilitate cargo recruitment and clathrin-coated pit nucleation. The prototypical clathrin adaptor AP2 mediates cargo recruitment, maturation, and scission of the pit by binding cargo, clathrin, and accessory proteins, including the Eps-homology (EH) domain proteins Eps15 and intersectin. However, clathrin-mediated endocytosis of some cargoes proceeds efficiently in AP2-depleted cells. We found that Dab2, another endocytic adaptor, also binds to Eps15 and intersectin. Depletion of EH domain proteins altered the number and size of clathrin structures and impaired the endocytosis of the Dab2- and AP2-dependent cargoes, integrin β1 and transferrin receptor, respectively. To test the importance of Dab2 binding to EH domain proteins for endocytosis, we mutated the EH domain-binding sites. This mutant localized to clathrin structures with integrin β1, AP2, and reduced amounts of Eps15. Of interest, although integrin β1 endocytosis was impaired, transferrin receptor internalization was unaffected. Surprisingly, whereas clathrin structures contain both Dab2 and AP2, integrin β1 and transferrin localize in separate pits. These data suggest that Dab2-mediated recruitment of EH domain proteins selectively drives the internalization of the Dab2 cargo, integrin β1. We propose that adaptors may need to be bound to their cargo to regulate EH domain proteins and internalize efficiently.
The Rho GTPases RhoA, Rac1, and Cdc42 play a major role in regulating the reorganization of the actin cytoskeleton. We recently identified CdGAP, a novel GTPase-activating protein with activity toward Rac1 and Cdc42. CdGAP consists of a N-terminal GAP domain, a central domain, and a C-terminal proline-rich domain. Here we show that through a subset of its Src homology 3 domains, the endocytic protein intersectin interacts with CdGAP. In platelet-derived growth factor-stimulated Swiss 3T3 cells, intersectin co-localizes with CdGAP and inhibits its GAP activity toward Rac1. Intersectin-Src homology 3 also inhibits CdGAP activity in GAP assays in vitro. Although the C-terminal proline-rich domain of CdGAP is required for the regulation of its GAP activity by intersectin both in vivo and in vitro, it is not necessary for CdGAP-intersectin interaction. Our data suggest that the central domain of CdGAP is required for CdGAP-intersectin interaction. Thus, we propose a model in which intersectin binding results in a change of CdGAP conformation involving the proline-rich domain that leads to the inhibition of its GAP activity. These observations provide the first demonstration of a direct regulation of RhoGAP activity through a protein-protein interaction and suggest a function for intersectin in Rac1 regulation and actin dynamics.
The Rho GTPases RhoA, Rac1, and Cdc42 play a major role in regulating the reorganization of the actin cytoskeleton. We recently identified CdGAP, a novel GTPase-activating protein with activity toward Rac1 and Cdc42. CdGAP consists of a N-terminal GAP domain, a central domain, and a C-terminal proline-rich domain. Here we show that through a subset of its Src homology 3 domains, the endocytic protein intersectin interacts with CdGAP. In platelet-derived growth factor-stimulated Swiss 3T3 cells, intersectin co-localizes with CdGAP and inhibits its GAP activity toward Rac1. Intersectin-Src homology 3 also inhibits CdGAP activity in GAP assays in vitro. Although the C-terminal proline-rich domain of CdGAP is required for the regulation of its GAP activity by intersectin both in vivo and in vitro, it is not necessary for CdGAP-intersectin interaction. Our data suggest that the central domain of CdGAP is required for CdGAP-intersectin interaction. Thus, we propose a model in which intersectin binding results in a change of CdGAP conformation involving the proline-rich domain that leads to the inhibition of its GAP activity. These observations provide the first demonstration of a direct regulation of RhoGAP activity through a protein-protein interaction and suggest a function for intersectin in Rac1 regulation and actin dynamics.
Endocytic adaptor proteins facilitate cargo recruitment and clathrin-coated pit nucleation. The prototypical clathrin adaptor AP2 mediates cargo recruitment, maturation, and scission of the pit by binding cargo, clathrin, and accessory proteins, including the Eps-homology (EH) domain proteins Eps15 and intersectin. However, clathrin-mediated endocytosis of some cargoes proceeds efficiently in AP2-depleted cells. We found that Dab2, another endocytic adaptor, also binds to Eps15 and intersectin. Depletion of EH domain proteins altered the number and size of clathrin structures and impaired the endocytosis of the Dab2- and AP2-dependent cargoes, integrin β1 and transferrin receptor, respectively. To test the importance of Dab2 binding to EH domain proteins for endocytosis, we mutated the EH domain-binding sites. This mutant localized to clathrin structures with integrin β1, AP2, and reduced amounts of Eps15. Of interest, although integrin β1 endocytosis was impaired, transferrin receptor internalization was unaffected. Surprisingly, whereas clathrin structures contain both Dab2 and AP2, integrin β1 and transferrin localize in separate pits. These data suggest that Dab2-mediated recruitment of EH domain proteins selectively drives the internalization of the Dab2 cargo, integrin β1. We propose that adaptors may need to be bound to their cargo to regulate EH domain proteins and internalize efficiently.
Using selected trapped exons with homology to specific protein domains, we identified a new full-length cDNA encoding a protein containing many motifs for protein-protein interactions. There are two major mRNA transcripts, a ubiquitously expressed mRNA of 5.3 kb and a brain-specific transcript of approximately 15 kb, encoding proteins of 1220 and 1721 amino acids, respectively. The stop codon of the ORF of the shorter transcript is split between adjacent exons. In brain tissues the last exon of the short transcript is skipped, and an alternative downstream exon, the first of several additional, is used to produce the 15-kb mRNA. The putative human protein is highly homologous to Xenopus intersectin (81% identical) and to Drosophila dynamin-associated protein, Dap160 (31% identical) and was termed intersectin (ITSN). Both human proteins contain five SH3 (Src homology 3) domains, two EH (Eps15 homology) domains, and an alpha-helix-forming region. The brain-specific long transcript encodes for three additional domains: a GEF (guanine-nucleotide exchange factors), a PH (pleckstrin homology), and a C2 domain. The Drosophila homologue is associated with dynamin, a protein family involved in the endocytic pathway and/or synaptic vesicle recycling. The structure of the human ITSN protein is consistent with its involvement in membrane-associated molecular trafficking and signal transduction pathways. The human ITSN gene has been mapped to 21q22. 1-q22.2 between markers D21S319 and D21S65, and its importance in Down syndrome and monogenic disorders is currently unknown.
Stimulates the exchange of guanyl nucleotides associated with a GTPase. Under normal cellular physiological conditions, the concentration of GTP is higher than that of GDP, favoring the replacement of GDP by GTP in association with the GTPase.
Using selected trapped exons with homology to specific protein domains, we identified a new full-length cDNA encoding a protein containing many motifs for protein-protein interactions. There are two major mRNA transcripts, a ubiquitously expressed mRNA of 5.3 kb and a brain-specific transcript of approximately 15 kb, encoding proteins of 1220 and 1721 amino acids, respectively. The stop codon of the ORF of the shorter transcript is split between adjacent exons. In brain tissues the last exon of the short transcript is skipped, and an alternative downstream exon, the first of several additional, is used to produce the 15-kb mRNA. The putative human protein is highly homologous to Xenopus intersectin (81% identical) and to Drosophila dynamin-associated protein, Dap160 (31% identical) and was termed intersectin (ITSN). Both human proteins contain five SH3 (Src homology 3) domains, two EH (Eps15 homology) domains, and an alpha-helix-forming region. The brain-specific long transcript encodes for three additional domains: a GEF (guanine-nucleotide exchange factors), a PH (pleckstrin homology), and a C2 domain. The Drosophila homologue is associated with dynamin, a protein family involved in the endocytic pathway and/or synaptic vesicle recycling. The structure of the human ITSN protein is consistent with its involvement in membrane-associated molecular trafficking and signal transduction pathways. The human ITSN gene has been mapped to 21q22. 1-q22.2 between markers D21S319 and D21S65, and its importance in Down syndrome and monogenic disorders is currently unknown.
Interacting selectively and non-covalently with a proline-rich region, i.e. a region that contains a high proportion of proline residues, in a protein.
Evidence
1:
Inferred from Physical InteractionUniProtKB
Intersectin 1 (ITSN1) is an evolutionarily conserved adaptor protein involved in clathrin-mediated endocytosis, cellular signaling and cytoskeleton rearrangement. ITSN1 gene is located on human chromosome 21 in Down syndrome critical region. Several studies confirmed role of ITSN1 in Down syndrome phenotype. Here we report the identification of novel interconnections in the interaction network of this endocytic adaptor. We show that the membrane-deforming protein SGIP1 (Src homology 3-domain growth factor receptor-bound 2-like (endophilin) interacting protein 1) and the signaling adaptor Reps1 (RalBP associated Eps15-homology domain protein) interact with ITSN1 in vivo. Both interactions are mediated by the SH3 domains of ITSN1 and proline-rich motifs of protein partners. Moreover complexes comprising SGIP1, Reps1 and ITSN1 have been identified. We also identified new interactions between SGIP1, Reps1 and the BAR (Bin/amphiphysin/Rvs) domain-containing protein amphiphysin 1. Immunofluorescent data have demonstrated colocalization of ITSN1 with the newly identified protein partners in clathrin-coated pits. These findings expand the role of ITSN1 as a scaffolding molecule bringing together components of endocytic complexes.
Interacting selectively and non-covalently with any protein or protein complex (a complex of two or more proteins that may include other nonprotein molecules).
Evidence
1:
Inferred from Physical InteractionIntAct
Disrupted-In-Schizophrenia 1 (DISC1) is a novel gene associated with schizophrenia by multiple genetic studies. In order to determine how mutations in DISC1 might cause susceptibility to schizophrenia, we undertook a comprehensive study of the cellular biology of DISC1 in its full-length and disease-associated mutant forms. DISC1 interacts by yeast two-hybrid, mammalian two-hybrid, and co-immunoprecipitation assays with multiple proteins of the centrosome and cytoskeletal system, including MIPT3, MAP1A and NUDEL; proteins which localize receptors to membranes, including alpha-actinin2 and beta4-spectrin; and proteins which transduce signals from membrane receptors, including ATF4 and ATF5. Truncated mutant DISC1 fails to interact with ATF4, ATF5 or NUDEL. Deletion mapping demonstrated that DISC1 has distinct interaction domains: MAP1A interacts via its LC2 domain with the N-terminus of DISC1, whereas MIPT3 and NUDEL bind via their C-terminal domains to the central coiled-coil domain of DISC1, and ATF4/5 bind via their C-terminal domains to the C-terminus of DISC1. In its full-length form, DISC1 protein localizes to predominantly perinuclear punctate structures which extend into neurites in some cells; mutant truncated DISC1, by contrast, is seen in a diffuse pattern throughout the cytoplasm and abundantly in neurites. Both forms co-localize with the centrosomal complex, although truncated less abundantly than full-length DISC1. Although both full-length and mutant DISC1 are found in microtubule fractions, neither form of DISC1 appears to bind directly to microtubules, but rather do so in a MIPT3-dependent fashion that is stabilized by taxol. Based on these data, we propose that DISC1 is a multifunctional protein whose truncation contributes to schizophrenia susceptibility by disrupting intracellular transport, neurite architecture and/or neuronal migration, all of which have been hypothesized to be pathogenic in the schizophrenic brain.
Evidence
2:
Inferred from Physical InteractionIntAct
Clathrin-mediated endocytosis, the major pathway for ligand internalization into eukaryotic cells, is thought to be initiated by the clustering of clathrin and adaptors around receptors destined for internalization. However, here we report that the membrane-sculpting F-BAR domain-containing Fer/Cip4 homology domain-only proteins 1 and 2 (FCHo1/2) were required for plasma membrane clathrin-coated vesicle (CCV) budding and marked sites of CCV formation. Changes in FCHo1/2 expression levels correlated directly with numbers of CCV budding events, ligand endocytosis, and synaptic vesicle marker recycling. FCHo1/2 proteins bound specifically to the plasma membrane and recruited the scaffold proteins eps15 and intersectin, which in turn engaged the adaptor complex AP2. The FCHo F-BAR membrane-bending activity was required, leading to the proposal that FCHo1/2 sculpt the initial bud site and recruit the clathrin machinery for CCV formation.
Evidence
3:
Inferred from Physical InteractionUniProtKB
Intersectin 1 (ITSN1) is an evolutionarily conserved adaptor protein involved in clathrin-mediated endocytosis, cellular signaling and cytoskeleton rearrangement. ITSN1 gene is located on human chromosome 21 in Down syndrome critical region. Several studies confirmed role of ITSN1 in Down syndrome phenotype. Here we report the identification of novel interconnections in the interaction network of this endocytic adaptor. We show that the membrane-deforming protein SGIP1 (Src homology 3-domain growth factor receptor-bound 2-like (endophilin) interacting protein 1) and the signaling adaptor Reps1 (RalBP associated Eps15-homology domain protein) interact with ITSN1 in vivo. Both interactions are mediated by the SH3 domains of ITSN1 and proline-rich motifs of protein partners. Moreover complexes comprising SGIP1, Reps1 and ITSN1 have been identified. We also identified new interactions between SGIP1, Reps1 and the BAR (Bin/amphiphysin/Rvs) domain-containing protein amphiphysin 1. Immunofluorescent data have demonstrated colocalization of ITSN1 with the newly identified protein partners in clathrin-coated pits. These findings expand the role of ITSN1 as a scaffolding molecule bringing together components of endocytic complexes.
Evidence
4:
Inferred from Physical InteractionIntAct
Activation of Rho-family GTPases involves the removal of bound GDP and the subsequent loading of GTP, all catalyzed by guanine nucleotide exchange factors (GEFs) of the Dbl-family. Despite high sequence conservation among Rho GTPases, Dbl proteins possess a wide spectrum of discriminatory potentials for Rho-family members. To rationalize this specificity, we have determined crystal structures of the conserved, catalytic fragments (Dbl and pleckstrin homology domains) of the exchange factors intersectin and Dbs in complex with their cognate GTPases, Cdc42 and RhoA, respectively. Structure-based mutagenesis of intersectin and Dbs reveals the key determinants responsible for promoting exchange activity in Cdc42, Rac1 and RhoA. These findings provide critical insight into the structural features necessary for the proper pairing of Dbl-exchange factors with Rho GTPases and now allow for the detailed manipulation of signaling pathways mediated by these oncoproteins in vivo.
Intersectin 1 (ITSN1) is an evolutionarily conserved adaptor protein involved in clathrin-mediated endocytosis, cellular signaling and cytoskeleton rearrangement. ITSN1 gene is located on human chromosome 21 in Down syndrome critical region. Several studies confirmed role of ITSN1 in Down syndrome phenotype. Here we report the identification of novel interconnections in the interaction network of this endocytic adaptor. We show that the membrane-deforming protein SGIP1 (Src homology 3-domain growth factor receptor-bound 2-like (endophilin) interacting protein 1) and the signaling adaptor Reps1 (RalBP associated Eps15-homology domain protein) interact with ITSN1 in vivo. Both interactions are mediated by the SH3 domains of ITSN1 and proline-rich motifs of protein partners. Moreover complexes comprising SGIP1, Reps1 and ITSN1 have been identified. We also identified new interactions between SGIP1, Reps1 and the BAR (Bin/amphiphysin/Rvs) domain-containing protein amphiphysin 1. Immunofluorescent data have demonstrated colocalization of ITSN1 with the newly identified protein partners in clathrin-coated pits. These findings expand the role of ITSN1 as a scaffolding molecule bringing together components of endocytic complexes.
Stimulates the exchange of guanyl nucleotides associated with a GTPase of the Rho family. Under normal cellular physiological conditions, the concentration of GTP is higher than that of GDP, favoring the replacement of GDP by GTP in association with the GTPase.
Any process that activates or increases the frequency, rate or extent of the protein kinase B signaling cascade, a series of reactions mediated by the intracellular serine/threonine kinase protein kinase B.
An endocytosis process that results in the invagination of the axonal plasma membrane to create a membrane-bounded vesicle. This process takes up excess membrane that would otherwise accumulate at the presynaptic terminal due to fusion of vesicle membranes during neurotransmitter release. The vesicles created may subsequently be used for neurotransmitter storage and release.
Using selected trapped exons with homology to specific protein domains, we identified a new full-length cDNA encoding a protein containing many motifs for protein-protein interactions. There are two major mRNA transcripts, a ubiquitously expressed mRNA of 5.3 kb and a brain-specific transcript of approximately 15 kb, encoding proteins of 1220 and 1721 amino acids, respectively. The stop codon of the ORF of the shorter transcript is split between adjacent exons. In brain tissues the last exon of the short transcript is skipped, and an alternative downstream exon, the first of several additional, is used to produce the 15-kb mRNA. The putative human protein is highly homologous to Xenopus intersectin (81% identical) and to Drosophila dynamin-associated protein, Dap160 (31% identical) and was termed intersectin (ITSN). Both human proteins contain five SH3 (Src homology 3) domains, two EH (Eps15 homology) domains, and an alpha-helix-forming region. The brain-specific long transcript encodes for three additional domains: a GEF (guanine-nucleotide exchange factors), a PH (pleckstrin homology), and a C2 domain. The Drosophila homologue is associated with dynamin, a protein family involved in the endocytic pathway and/or synaptic vesicle recycling. The structure of the human ITSN protein is consistent with its involvement in membrane-associated molecular trafficking and signal transduction pathways. The human ITSN gene has been mapped to 21q22. 1-q22.2 between markers D21S319 and D21S65, and its importance in Down syndrome and monogenic disorders is currently unknown.
Studies in clathrin-mediated endocytosis of ITGB1 and TFR used a siRNA mixture of ISTN1 and ISTN2, and a Dab2 mutant with impaired binding to EH domain-containing proteins EPS15 and ITSN1 suggesting a partially overlapping role of the EH domain-containing proteins (PubMed22648170).
Endocytic adaptor proteins facilitate cargo recruitment and clathrin-coated pit nucleation. The prototypical clathrin adaptor AP2 mediates cargo recruitment, maturation, and scission of the pit by binding cargo, clathrin, and accessory proteins, including the Eps-homology (EH) domain proteins Eps15 and intersectin. However, clathrin-mediated endocytosis of some cargoes proceeds efficiently in AP2-depleted cells. We found that Dab2, another endocytic adaptor, also binds to Eps15 and intersectin. Depletion of EH domain proteins altered the number and size of clathrin structures and impaired the endocytosis of the Dab2- and AP2-dependent cargoes, integrin β1 and transferrin receptor, respectively. To test the importance of Dab2 binding to EH domain proteins for endocytosis, we mutated the EH domain-binding sites. This mutant localized to clathrin structures with integrin β1, AP2, and reduced amounts of Eps15. Of interest, although integrin β1 endocytosis was impaired, transferrin receptor internalization was unaffected. Surprisingly, whereas clathrin structures contain both Dab2 and AP2, integrin β1 and transferrin localize in separate pits. These data suggest that Dab2-mediated recruitment of EH domain proteins selectively drives the internalization of the Dab2 cargo, integrin β1. We propose that adaptors may need to be bound to their cargo to regulate EH domain proteins and internalize efficiently.
Protein involved in endocytosis, a process by which extracellular materials are taken up into a cell by invagination of the plasma membrane to form vesicles enclosing these materials.
A reference proteome is a set of protein sequences derived from a complete proteome which constitutes a defined standard for a particular user community. Reference proteomes are manually defined according to a number of criteria. They cover the proteomes of well- studied model organisms and other proteomes of interest for biomedical and biotechnological research. Reference proteomes have been selected to provide broad coverage of the tree of life, and constitute a representative cross-section of the taxonomic diversity to be found within UniProtKB.
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