Identification of Rab GTPase-Activating Protein-Like Protein (RabGAPLP) as a Novel Alix/AIP1-Interacting Protein

Ichioka, Fumitaka; Horii, Mio; Katoh, Keiichi; Terasawa, Yoshinori; Shibata, Hideki; Maki, Masatoshi

doi:10.1271/bbb.69.861

Cited by 8 publications

(10 citation statements)

References 20 publications

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“…To test this hypothesis, we examined the ability of Bro i to bind the three isoforms of CHMP4 using immunoprecipitation assays. Although Bro i carries part of the CHMP4 binding interface [44],[46],[47],[66], it was unable to interact with the three CHMP4 isoforms (Figure S3A, S3B, and S3C), even though it retained binding to MAP/RabGAPLP, a known cellular partner of the Bro1 domain of Alix [67] (Figure S3D). The inability of Bro i to bind CHMP4 isoforms was also observed in yeast two hybrid assays (data not shown).…”

Section: Resultsmentioning

confidence: 99%

The Nucleocapsid Region of HIV-1 Gag Cooperates with the PTAP and LYPXnL Late Domains to Recruit the Cellular Machinery Necessary for Viral Budding

et al. 2009

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HIV-1 release is mediated through two motifs in the p6 region of Gag, PTAP and LYPXnL, which recruit cellular proteins Tsg101 and Alix, respectively. The Nucleocapsid region of Gag (NC), which binds the Bro1 domain of Alix, also plays an important role in HIV-1 release, but the underlying mechanism remains unclear. Here we show that the first 202 residues of the Bro1 domain (Broi) are sufficient to bind Gag. Broi interferes with HIV-1 release in an NC–dependent manner and arrests viral budding at the plasma membrane. Similar interrupted budding structures are seen following over-expression of a fragment containing Bro1 with the adjacent V domain (Bro1-V). Although only Bro1-V contains binding determinants for CHMP4, both Broi and Bro1-V inhibited release via both the PTAP/Tsg101 and the LYPXnL/Alix pathways, suggesting that they interfere with a key step in HIV-1 release. Remarkably, we found that over-expression of Bro1 rescued the release of HIV-1 lacking both L domains. This rescue required the N-terminal region of the NC domain in Gag and the CHMP4 binding site in Bro1. Interestingly, release defects due to mutations in NC that prevented Bro1 mediated rescue of virus egress were rescued by providing a link to the ESCRT machinery via Nedd4.2s over-expression. Our data support a model in which NC cooperates with PTAP in the recruitment of cellular proteins necessary for its L domain activity and binds the Bro1–CHMP4 complex required for LYPXnL–mediated budding.

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Section: Resultsmentioning

confidence: 99%

The Nucleocapsid Region of HIV-1 Gag Cooperates with the PTAP and LYPXnL Late Domains to Recruit the Cellular Machinery Necessary for Viral Budding

et al. 2009

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“…Alix functions as a scaffold protein in multiple cellular processes (Carlton and Martin-Serrano, 2007; Ichioka et al, 2005; Katoh et al, 2003; Mahul-Mellier et al, 2008; Martin-Serrano et al, 2003a; Matsuo et al, 2004; Missotten et al, 1999; Morita et al, 2007a; Odorizzi, 2006; Odorizzi et al, 2003; Pan et al, 2008; Pan et al, 2006; Sadoul, 2006; Sette et al, 2010; Vito et al, 1999; Zhou et al, 2010) and appears to fulfill a similar function in HIV-1 release. It binds Gag during virus assembly (Jouvenet et al, 2011) and is believed to play a key role in the recruitment of activated CHMP4 isoforms at the budding neck (Fisher et al, 2007; Pires et al, 2009; Strack et al, 2003; Usami et al, 2007).…”

Section: Discussionmentioning

confidence: 99%

“…In compliance with such a role, comparison of the crystal structures of the Bro1 domain of Alix, HD-PTP and Brox (Figure 2) identified an additional docking region in the Bro1 domain of Alix at the tip of the Phe105 loop. The N-terminal 423 residues of Alix, but not those from HD-PTP or Brox, have been shown to bind the C-terminal fragment of RabGAPLP or RabGAP-5 containing SH3 and RUN domains (Haas et al, 2005; Ichioka et al, 2005). RUN domains are commonly involved in regulation of small GTPase functions (Callebaut et al, 2001; Haas et al, 2005; Ichioka et al, 2005).…”

Section: Discussionmentioning

confidence: 99%

The Phe105 Loop of Alix Bro1 Domain Plays a Key Role in HIV-1 Release

Sette

Dussupt

et al. 2011

Structure

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Summary Alix and cellular paralogs HD-PTP and Brox contain N-terminal Bro1 domains that bind ESCRT-III CHMP4. In contrast to HD-PTP and Brox, expression of the Bro1 domain of Alix alleviates HIV-1 release defects due to interrupted access to ESCRT. In an attempt to elucidate this functional discrepancy, we solved the crystal structures of the Bro1 domains of HD-PTP and Brox. They revealed typical “boomerang” folds they share with the Bro1 Alix domain. However, they each contain unique structural features that may be relevant to their specific function(s). In particular, phenylalanine residue in position 105 (Phe105) of Alix belongs to a long loop that is unique to its Bro1 domain. Concurrently mutation of Phe105 and surrounding residues at the tip of the loop compromises the function of Alix in HIV-1 budding without affecting its interactions with Gag or CHMP4. These studies identify a new functional determinant in the Bro1 domain of Alix.

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“…These functions include lysobisphosphatidic acid (LBPA) binding (reviewed in ref. 20), endophilin binding (21), receptor trafficking (22)(23)(24), endosome distribution (25), cell motility/adhesion (26,27), apoptosis (reviewed in ref. 28), actin and microtubule binding (26,29) and regulation of JNK signaling (30).…”

mentioning

confidence: 99%

ALIX-CHMP4 interactions in the human ESCRT pathway

McCullough

Fisher

Whitby

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

214

252

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The ESCRT pathway facilitates membrane fission events during enveloped virus budding, multivesicular body formation, and cytokinesis. To promote HIV budding and cytokinesis, the ALIX protein must bind and recruit CHMP4 subunits of the ESCRT-III complex, which in turn participate in essential membrane remodeling functions. Here, we report that the Bro1 domain of ALIX binds specifically to C-terminal residues of the human CHMP4 proteins (CHMP4A-C). Crystal structures of the complexes reveal that the CHMP4 C-terminal peptides form amphipathic helices that bind across the conserved concave surface of ALIXBro1. ALIX-dependent HIV-1 budding is blocked by mutations in exposed ALIXBro1 residues that help contribute to the binding sites for three essential hydrophobic residues that are displayed on one side of the CHMP4 recognition helix (M/L/IxxLxxW). The homologous CHMP1-3 classes of ESCRT-III proteins also have C-terminal amphipathic helices, but, in those cases, the three hydrophobic residues are arrayed with L/I/MxxxLxxL spacing. Thus, the distinct patterns of hydrophobic residues provide a ''code'' that allows the different ESCRT-III subunits to bind different ESCRT pathway partners, with CHMP1-3 proteins binding MIT domain-containing proteins, such as VPS4 and Vta1/LIP5, and CHMP4 proteins binding Bro1 domaincontaining proteins, such as ALIX.cytokinesis ͉ ESCRT-III ͉ HIV ͉ mutivesicular body

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Identification of Rab GTPase-Activating Protein-Like Protein (RabGAPLP) as a Novel Alix/AIP1-Interacting Protein

Cited by 8 publications

References 20 publications

The Nucleocapsid Region of HIV-1 Gag Cooperates with the PTAP and LYPXnL Late Domains to Recruit the Cellular Machinery Necessary for Viral Budding

The Nucleocapsid Region of HIV-1 Gag Cooperates with the PTAP and LYPXnL Late Domains to Recruit the Cellular Machinery Necessary for Viral Budding

The Phe105 Loop of Alix Bro1 Domain Plays a Key Role in HIV-1 Release

ALIX-CHMP4 interactions in the human ESCRT pathway

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