Tom1 (target of Myb 1) and its related proteins (Tom1L1 and Tom1L2) constitute a protein family and share an N-terminal VHS (Vps27p/Hrs/Stam) domain followed by a GAT (GGA and Tom1) domain.VHS domains are found at the N termini of select proteins involved in intracellular membrane trafficking and are often localized to membranes. The three dimensional structure of human TOM1 VHS domain reveals eight helices arranged in a superhelix. The surface of the domain has two main features: (1) a basic patch on one side due to several conserved positively charged residues on helix 3 and (2) a negatively charged ridge on the opposite side, formed by residues on helix 2 []. The basic patch is thought to mediate membrane binding.It was demonstrated that the GAT domain of both Tom1 and Tom1L1 binds ubiquitin, suggesting that these proteins might participate in the sorting of ubiquitinated proteins into multivesicular bodies (MVB) []. Moreover, Tom1L1 interacts with members of the MVB sorting machinery. Specifically, the VHS domain of Tom1L1 interacts with Hrs (hepatocyte growth factor-regulated tyrosine kinase substrate), whereas a PTAP motif, located between the VHS and GAT domains of Tom1L1, is responsible for binding to TSG101 (tumour susceptibility gene 101). Myc epitope-tagged Tom1L1 is recruited to endosomes following Hrs expression. In addition, Tom1L1 possesses several tyrosine motifs at the C-terminal region that mediate interactions with members of the Src family kinases and other signalling proteins such as Grb2 and p85. Expression of a constitutively active form of Fyn kinase promotes the recruitment of Tom1L1 to enlarged endosomes. It is proposed that Tom1L1 could act as an intermediary between the signalling and degradative pathways [].Over expression of Tom1 suppresses activation of the transcription factors NF-kappaB and AP-1, induced by either IL-1beta or tumour necrosis factor (TNF)-alpha, and the VHS domain of Tom1 is indispensable for this suppressive activity. This suggests that Tom1 is a common negative regulator of signalling pathways induced by IL-1beta and TNF-alpha [].
This domain is found in plant proteins including THH1/TOM1/TOM3 from Arabidopsis. TOM1 and TOM3 are transmembrane proteins necessary for the efficient multiplication of tobamoviruses []. THH1 supports tobamovirus multiplication, but to a lesser extent than TOM1 and TOM3 []. Members containing this domain are part of the GPCR superfamily and involved in stress tolerance [, ].
This entry represents a group of plant proteins, including THH1/TOM1/TOM3 from Arabidopsis. TOM1 and TOM3 are transmembrane proteins necessary for the efficient multiplication of tobamoviruses []. THH1 supports tobamovirus multiplication, but to a lesser extent than TOM1 and TOM3 []. Proteins in this family belong to the GCPR superfamily and are also involved in stress tolerance [, ].
This entry includes TOM1-like protein 1-9 (TOL1-9) from Arabidopsis [, ]. In general, they contain the VHS and GAT domains that bind to phospholipids and ubiquitin []. They may be part of the ESCRT (Endosomal Sorting Complex Required for Transport) complex contributing to the sorting of ubiquinated proteins to the MVB (multivesicular body) formation machinery. In plants they play a role in the control of plant morphogenesis []. This entry also includes TOM1 from Dictyostelium discoideum [].
Tollip (Toll-interacting protein) is a component of the IL-1RI pathway which contains an N-terminal C2 domain and a C-terminal CUE domain. Tollip binds to the cytoplasmic TIR domain of IL-1Rs after IL-1 stimulation. It is sufficient for recruitment of IRAK to IL-1Rs and negatively regulates IL-1-induced signaling by inhibiting IRAK phosphorylation. In addition, Tollip directly interacts with toll-like receptors TLR2 and TLR4, and plays an inhibitory role in TLR-mediated cell activation through suppressing phosphorylation and kinase activity of IRAK. Moreover, Tollip can associate with GAT domains of Tom1 and its related proteins Tom1L1 and Tom1L2, and facilitate the recruitment of clathrin onto endosomes [, ].
The VHS domain is an about 150 residues long domain, whose name is derivedfrom its occurrence in VPS-27, Hrs and STAM. The VHS domain is found at the N-termini of proteins associated with endocytocis and/or vesicular trafficking,often in association with other domains like FYVE, SH3 or TAM [, ]. The VHS domain of Hrs makes both intra- andintermolecular interactions with FYVE domains and it has been proposed that itmight as well interact with other domains. The VHS domain might function as amultipurpose docking adapter that localizes proteins to the membrane throughinteractions with the membrane and/or the endocytic machinery [, ].Resolution of the crystal structure of the VHS domain of Drosophila Hrs andhuman Tom1 revealed that it consists of eight helices arranged in a superhelix[, ].
This domain is found in the N-terminal region of the GGA and Tom1 (GAT) domain in Golgi-localizing gamma-adaptin ARF-binding protein 1 (GGA1). This domain can also be found in GGA2 and GGA3. The GAT domains is the key region in GGA that interacts with ARF. ARF plays a crucial role in docking adaptor proteins to membranes. This domain is referred to as N-GAT and it interacts extensively with ARF [].The GAT domain is a region of homology of ~130 residues, which is found in eukaryotic GGAs (for Golgi-localized, gamma ear-containing ADP ribosylation factor (ARF)-binding proteins) and vertebrate TOMs (for target of myb). The GAT domain is found in its entirety only in GGAs, although, at the C terminus it shares partial sequence similarity with a short region of TOMs. The GAT domain is found in association with other domains, such as VHS and GAE. The GAT domain of GGAs serves as a molecular anchor of GGA to trans-Golgi network (TGN) membranes via its interaction with the GTP-bound form of a member of the ARFfamily of small GTPases and can bind specifically to the Rab GTPase effector rabaptin5 and to ubiquitin [, , , ].The GGA-GAT domain possesses an all α-helical structure, composed of four helices arranged in a somewhat unusual topology, which has been called the helical paper clip. The overall structure shows that the GAT domain has an elongated shape, in which the longest helix participates in two small independent subdomains: an N-terminal helix-loop-helix hook and a C-terminal three-helix bundle. The hook subdomain has been shown to be both necessary and sufficient for ARF-GTP binding and Golgi targeting of GGAs. The N-terminal hook subdomain contains a hydrophobic patch, which is found to interact directly with ARF []. It has been proposed that this interaction might stabilise the hook subdomain []. The C-terminal three-helix bundle is involved in the binding with Rabaptin5 and ubiquitin [].
Proteins synthesized on the ribosome and processed in the endoplasmic reticulum are transported from the Golgi apparatus to the trans-Golgi network (TGN), and from there via small carrier vesicles to their final destination compartment. These vesicles have specific coat proteins (such as clathrin or coatomer) that are important for cargo selection and direction of transport []. Clathrin coats contain both clathrin (acts as a scaffold) and adaptor complexes that link clathrin to receptors in coated vesicles. Clathrin-associated protein complexes are believed to interact with the cytoplasmic tails of membrane proteins, leading to their selection and concentration. The two major types of clathrin adaptor complexes are the heterotetrameric adaptor protein (AP) complexes, and the monomeric GGA (Golgi-localising, Gamma-adaptin ear domain homology, ARF-binding proteins) adaptors [, ].AP (adaptor protein) complexes are found in coated vesicles and clathrin-coated pits. AP complexes connect cargo proteins and lipids to clathrin at vesicle budding sites, as well as binding accessory proteins that regulate coat assembly and disassembly (such as AP180, epsins and auxilin). There are different AP complexes in mammals. AP1 is responsible for the transport of lysosomal hydrolases between the TGN and endosomes []. AP2 associates with the plasma membrane and is responsible for endocytosis []. AP3 is responsible for protein trafficking to lysosomes and other related organelles []. AP4 is less well characterised. AP complexes are heterotetramers composed of two large subunits (adaptins), a medium subunit (mu) and a small subunit (sigma). For example, in AP1 these subunits are gamma-1-adaptin, beta-1-adaptin, mu-1 and sigma-1, while in AP2 they are alpha-adaptin, beta-2-adaptin, mu-2 and sigma-2. Each subunit has a specific function. Adaptins recognise and bind to clathrin through their hinge region (clathrinbox), and recruit accessory proteins that modulate AP function through their C-terminal ear (appendage) domains. Mu recognises tyrosine-based sorting signals within the cytoplasmic domains of transmembrane cargo proteins []. One function of clathrin and AP2 complex-mediated endocytosis is to regulate the number of GABA(A) receptors available at the cell surface []. GGAs (Golgi-localising, Gamma-adaptin ear domain homology, ARF-binding proteins) are a family of monomeric clathrin adaptor proteins that are conserved from yeasts to humans. GGAs regulate clathrin-mediated the transport of proteins (such as mannose 6-phosphate receptors) from the TGN to endosomes and lysosomes through interactions with TGN-sorting receptors, sometimes in conjunction with AP-1 [, ]. GGAs bind cargo, membranes, clathrin and accessory factors. GGA1, GGA2 and GGA3 all contain a domain homologous to the ear domain of gamma-adaptin. GGAs are composed of a single polypeptide with four domains: an N-terminal VHS (Vps27p/Hrs/Stam) domain, a GAT (GGA and Tom1) domain, a hinge region, and a C-terminal GAE (gamma-adaptin ear) domain. The VHS domain is responsible for endocytosis and signal transduction, recognising transmembrane cargo through the ACLL sequence in the cytoplasmic domains of sorting receptors []. The GAT domain (also found in Tom1 proteins) interacts with ARF (ADP-ribosylation factor) to regulate membrane trafficking [], and with ubiquitin for receptor sorting []. The hinge region contains a clathrin box for recognition and binding to clathrin, similar to that found in AP adaptins. The GAE domain is similar to the AP gamma-adaptin ear domain, and is responsible for the recruitment of accessory proteins that regulate clathrin-mediated endocytosis [].This entry represents a β-sandwich structural motif found in the appendage (ear) domain of alpha-, beta- and gamma-adaptin from AP clathrin adaptor complexes, and the GAE (gamma-adaptin ear) domain of GGA adaptor proteins. These domains have an immunoglobulin-like β-sandwich fold containing 7 or 8 strands in 2 β-sheets in a Greek key topology [, ]. Although these domains share a similar fold, there is little sequence identity between the alpha/beta-adaptins and gamma-adaptin/GAE.
