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Review
. 2021 Jul 14;10(7):1780.
doi: 10.3390/cells10071780.

The Interactome of the VAP Family of Proteins: An Overview

Christina James  1 , Ralph H Kehlenbach  1
Affiliations
Review

The Interactome of the VAP Family of Proteins: An Overview

Christina James et al. Cells. .

Abstract

Membrane contact sites (MCS) are sites of close apposition of two organelles that help in lipid transport and synthesis, calcium homeostasis and several other biological processes. The VAMP-associated proteins (VAPs) VAPA, VAPB, MOSPD2 and the recently described MOSPD1 and MOSPD3 are tether proteins of MCSs that are mainly found at the endoplasmic reticulum (ER). VAPs interact with various proteins with a motif called FFAT (two phenylalanines in an acidic tract), recruiting the associated organelle to the ER. In addition to the conventional FFAT motif, the recently described FFNT (two phenylalanines in a neutral tract) and phospho-FFAT motifs contribute to the interaction with VAPs. In this review, we summarize and compare the recent interactome studies described for VAPs, including in silico and proximity labeling methods. Collectively, the interaction repertoire of VAPs is very diverse and highlights the complexity of interactions mediated by the different FFAT motifs to the VAPs.

Keywords: FFAT; MOSPD1; MOSPD2; MOSPD3; VAPA; VAPB; interactome.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Domain structures of the VAP family of proteins. All VAP family of proteins have an MSP (Major Sperm Protein) domain. Both VAPA and VAPB contain a central coiled-coil domain (CC) and a C-terminal transmembrane domain (TMD). VAPC lacks the CC domain and the TMD. In addition to the MSP domain, MOSPD1 and MOSPD3 contain two TMDs at the C-terminal end, whereas MOSPD2 possesses a CRAL/TRIO (cellular retinaldehyde-binding protein and triple functional domain protein) domain at the N-terminal and a TMD at the C-terminal end.
Figure 2
Figure 2
The structure of VAP-FFAT complexes. (A) Schematic representation of a conventional FFAT motif binding the MSP domain of VAP. The electropositive surface of the MSP domain binds the core residues of the FFAT motif, and the dotted lines represent the acidic tract. (B) The crystal structure of rat MSP-VAPA in complex with the ORP1 FFAT motif as described by Kaiser et al. [19] (PDB ID: 1Z9O). A tetramer formed by MSP-FFAT complex is shown. (B–D) The MSP domain is represented as ribbons (red: helix, yellow: β-sheet, green: loop), and the FFAT motif is represented as sticks (blue) that bind across the MSP domain. (C) The crystal structure of human MSP-VAPA in complex with OSBP-FFAT as described by Furuita et al. [51] (PDB ID: 2RR3). (D) Structure of human-MSP-VAPA in complex with STARD3 FFAT as described by Di Mattia et al. [52] (PDB ID: 6TQR).
Figure 3
Figure 3
Approaches used to analyze the interactome of the VAPs. (A) Schematic of high-throughput interaction mapping using affinity-purification mass spectrometry. From the human ORFeome, a lentiviral library was created. The baits were expressed in HEK293T cells, subjected to affinity purification, and protein complexes were analyzed by LC-MS. All the interactors and baits were combined to generate a network of the human interactome [43]. (B) In a BioID approach, the protein of interest is tagged with biotin ligase and expressed in cells. The enzyme converts free biotin into reactive biotinoyl-5′-AMP, which reacts with primary amines of proximal proteins. The biotinylated proteins are enriched and identified using mass spectrometry [59]. (C) In APEX-based proximity labelling, the protein of interest is fused to ascorbate peroxidase (APEX). In the presence of H2O2, APEX converts biotin phenol into biotin phenoxyl radicals, which covalently label proteins in close proximity. In RAPIDS, the specific subcellular interactome of VAPB was analyzed by using rapamycin induced dimerization of FKBP12- and FRB-containing fusion proteins, followed by APEX2-dependent biotinylation of proteins and their identification by MS [60]. In this approach, SILAC allows a direct comparison between plus- and minus-rapamycin conditions and, thus, a very efficient elimination of the background.
Figure 4
Figure 4
The interactome of VAPA and VAPB. (A) The Bioplex interaction network of both VAPA and VAPB in HEK293T cells reported by Huttlin et al. [43,57,58]. The proteins represented in blue are sole VAPA interactors, and those represented in green are sole VAPB interactors. The proteins represented in orange are common interacting partners of both VAPA and VAPB. (B) The Venn diagram shows VAPB interactors identified by Huttlin et al. (affinity purification-mass spectrometry) [43,57,58], by Cabukusta et al. (BioID followed by mass spectrometry identification) [22] and by James et al. (RAPIDS (rapamycin and apex dependent identification of proteins by SILAC) followed by mass spectrometry identification) [60]. See Table 1 for proteins that were identified in at least two studies.
Figure 5
Figure 5
Interactome of the MOSPD proteins. (A). The overlap of proteins interacting with MOSPD2 as identified by Di Mattia et al. [21] and Cabukusta et al. [22]. The overlapping proteins are RAB3GAP1/2, CCDC47, RMDN3, ACSL3, FAF2 and TMPO. (B) Venn diagram showing the overlap of interacting partners of all five proteins of the VAP family. The proteins interacting with MOSPD2, MOSPD1, MOSPD3 and/or VAPA/VAPB were identified by Di Mattia et al. [21], Cabukusta et al. [22], Huttlin et al. [43,57,58], James et al. [60], as indicated. The seven proteins shown to interact with all proteins are TACC1, FNDC3A, ESYT1, ESYT2, ANKLE2, ZC3HAV1 and EIF2AK3.
Figure 6
Figure 6
Proteins of the VAP family connect the ER and other organelles. Schematic representation of the interaction of all the family members with FFAT-motif at MCSs of the ER and endosomes, mitochondria, the plasma membrane, the Golgi apparatus, peroxisomes and the nuclear envelope. VAP proteins and a few examples of interacting proteins (FFAT-motif containing proteins) at the MCSs are depicted close to the organelles.
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