A Family of ADP-Ribosylation Factor Effectors That Can Alter Membrane Transport through thetrans-Golgi

Abstract

A family of three structurally related proteins were cloned from human cDNA libraries by their ability to interact preferentially with the activated form of human ADP-ribosylation factor 3 (ARF3) in two-hybrid assays. The specific and GTP-dependent binding was later confirmed through direct protein binding of recombinant proteins. The three proteins share large (Ϸ300 residues) domains at their N termini that are 60 -70% identical to each other and a shorter (73 residues) domain at their C termini with 70% homology to the C-terminal "ear" domain of ␥-adaptin. Although GGA1 is found predominantly as a soluble protein by cell fractionation, all three proteins were found to localize to the trans-Golgi network (TGN) by indirect immunofluorescence. The binding of GGAs to TGN was sensitive to brefeldin A, consistent with this being an ARF-dependent event. Thus, these proteins have been named Golgi-localizing, ␥-adaptin ear homology domain, ARF-binding proteins, or GGAs. The finding that overexpression of GGAs was sufficient to alter the distribution of markers of the TGN (TGN38 and mannose 6-phosphate receptors) led us to propose that GGAs are effectors for ARFs that function in the regulation of membrane traffic through the TGN.

Interactions of TOM1L1 with the Multivesicular Body Sorting Machinery

“…GGA proteins are monomeric adaptors that are recruited to the TGN by the Arf-1 GTPase (22)(23)(24)(25) and mediate the transport of lysosomal enzymes. They consist of four distinct segments as follows: a VHS domain that binds an acidic di-leucine sorting signal found in the mannose 6-phosphate receptor and other proteins known to cycle between TGN and endosomes (26,27); a GAT (GGA and Tom) domain that interacts with the GTPbound form of Arf (22)(23)28); a hinge region that recruits clathrin (27,28); and a similar to the ␥-adaptin ear domain that exhibits sequence similarity to the ear region of ␥-adaptin and recruits a number of accessory proteins (24, 29 -31). Most interestingly, we have recently described (18) that GGAs are present not only at the TGN but also at endosomes.…”

Autoinhibition of the ligand-binding site of GGA1/3 VHS domains by an internal acidic cluster-dileucine motif

“…T he GGAs (Golgi-localizing, ␥-adaptin ear homology domain, ARF-binding proteins) are a recently described family of proteins involved in trafficking between the Golgi and endosomes (1)(2)(3)(4)(5). Three GGAs have been identified in humans (GGA1, GGA2, and GGA3).…”

“…Three GGAs have been identified in humans (GGA1, GGA2, and GGA3). These proteins have modular structures consisting of an N-terminal VHS (VPS-27, Hrs, and STAM) domain followed by a GAT (GGA and TOM1) domain, a connecting hinge segment, and a C-terminal GAE domain, which has homology to the ear domain of ␥-adaptin (1)(2)(3)(4)(5). The VHS and GAT domains are highly conserved among the three mammalian GGAs with 60-75% identity across the N-terminal 300 aa (4).…”

“…These proteins have modular structures consisting of an N-terminal VHS (VPS-27, Hrs, and STAM) domain followed by a GAT (GGA and TOM1) domain, a connecting hinge segment, and a C-terminal GAE domain, which has homology to the ear domain of ␥-adaptin (1)(2)(3)(4)(5). The VHS and GAT domains are highly conserved among the three mammalian GGAs with 60-75% identity across the N-terminal 300 aa (4). The GAT domain binds to the GTP-bound form of the ADP-ribosylation factor (ARF) family of proteins that serve to recruit the GGAs from the cytosol to the Golgi complex (4,6,7).…”

“…The VHS and GAT domains are highly conserved among the three mammalian GGAs with 60-75% identity across the N-terminal 300 aa (4). The GAT domain binds to the GTP-bound form of the ADP-ribosylation factor (ARF) family of proteins that serve to recruit the GGAs from the cytosol to the Golgi complex (4,6,7). The VHS domain then interacts specifically with the acidic cluster (AC)-dileucine motif found in the cytoplasmic tails of sorting receptors such as sortilin, the cation-independent mannose 6-phosphate receptor (CI-MPR), and the cation-dependent mannose 6-phosphate receptor (CD-MPR), as well as the lowdensity lipoprotein receptor-related protein 3 (LRP3) (8)(9)(10)(11)(12)(13).…”