Phylogenetic Analysis of Sec7-Domain–containing Arf Nucleotide Exchangers

Abstract

The eukaryotic family of ADP-ribosylation factor (Arf) GTPases plays a key role in the regulation of protein trafficking, and guanine-nucleotide exchange is crucial for Arf function. Exchange is stimulated by members of another family of proteins characterized by a 200-amino acid Sec7 domain, which alone is sufficient to catalyze exchange on Arf. Here, we analyzed the phylogeny of Sec7-domain-containing proteins in seven model organisms, representing fungi, plants, and animals. The phylogenetic tree has seven main groups, of which two include members from all seven model systems. Three groups are specific for animals, whereas two are specific for fungi. Based on this grouping, we propose a phylogenetically consistent set of names for members of the Sec7-domain family. Each group, except for one, contains proteins with known Arf exchange activity, implying that all members of this family have this activity. Contrary to the current convention, the sensitivity of Arf exchange activity to the inhibitor brefeldin A probably cannot be predicted by group membership. Multiple alignment reveals group-specific domains outside the Sec7 domain and a set of highly conserved amino acids within it. Determination of the importance of these conserved elements in Arf exchange activity and other cellular functions is now possible.

Mutations in a Highly Conserved Region of the Arf1p ActivatorGEA2Block Anterograde Golgi Transport but Not COPI Recruitment to Membranes

“…A region of ϳ200 amino acids just upstream of the Sec7 domain had the next-highest frequency of mutations. This region contains residues evolutionarily conserved in Arf GEFs of both the Gea/GBF and Sec7/BIG subfamilies, and we refer to it as B/G␤ (Cox et al, 2004; Figure 2B). Of the 50 mutants with one or two amino acid substitutions, we tested 44 by Western blot analysis to determine the stability of the mutant protein.…”

“…Sec21p-GFP was present in a pattern of spots in all four mutants even after shift to the nonpermissive temperature for 40 min (Figure 4 and our unpublished data). For the wild-type GEA2 control and for each (Cox et al, 2004). The portion of the B/G␤ region (dark blue) that was indicated in A is shown in light blue, and its sequence, aligned with the homologous region from other Arf GEFs of the same subfamily, is shown.…”

“…The Golgi-localized Arf GEFs are large proteins (ϳ200 kDa) with multiple homology domains flanking the catalytic Sec7 domain (Jackson and Casanova, 2000;Cox et al, 2004). Three of these homology domains also share similarity to the BIG/Sec7 subfamily of Arf GEFs and are called B/G␣, B/G␤, and B/G␥ (Cox et al, 2004).…”

Dynamics of GBF1, a Brefeldin A-Sensitive Arf1 Exchange Factor at the Golgi

“…The Arf GEFs are responsible for the spatial and temporal regulation of Arf activation and all contain a centrally located Sec7 domain that is the catalytic domain for nucleotide exchange (Jackson and Casanova, 2000). There are six subfamilies of Arf GEFs in eukaryotic cells, one of which is the GEA/GBF subfamily, with homologues in all eukaryotes, including yeast, plants, and mammals (Cox et al, 2004). The Arf GEFs are all peripherally associated membrane proteins (Jackson and Casanova, 2000).…”

“…The lower molecular weight GEFs such as ARNO and the cytohesins possess a pleckstrin homology (PH) domain that mediates membrane binding through direct interaction with specific phosphoinositides (Cullen and Chardin, 2000). The high-molecular-weight Arf GEFs associated with the Golgi apparatus, GBF1, BIG1, and BIG2, do not have recognizable PH domains, and little is known about the mechanism of their interaction with membranes (Cox et al, 2004). Because the Arf GEFs determine when and where Arf proteins will be activated, it is critical that they be localized to the correct membrane site.…”