Interaction of the HIV-1 Intasome with Transportin 3 Protein (TNPO3 or TRN-SR2)

Larue, Ross C.; Gupta, Kushol; Wuensch, Christiane; Shkriabai, Nikolozi; Kessl, Jacques J.; Danhart, Eric; Feng, Lei; Taltynov, Oliver; Christ, Frauke; Duyne, Gregory D. Van; Debyser, Zeger; Foster, Mark P.; Kvaratskhelia, Mamuka

doi:10.1074/jbc.m112.384669

Cited by 52 publications

(67 citation statements)

References 83 publications

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“…S1A). Tnpo3 is known to dimerize at high protein concentrations, and the overall shape of the observed dimers is compatible with the molecular envelope determined by small-angle X-ray scattering (43).…”

Section: Resultssupporting

confidence: 48%

Structural basis for nuclear import of splicing factors by human Transportin 3

Maertens

Cook

Wang

et al. 2014

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

130

167

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Transportin 3 (Tnpo3, Transportin-SR2) is implicated in nuclear import of splicing factors and HIV-1 replication. Herein, we show that the majority of cellular Tnpo3 binding partners contain arginineserine (RS) repeat domains and present crystal structures of human Tnpo3 in its free as well as GTPase Ran-and alternative splicing factor/splicing factor 2 (ASF/SF2)-bound forms. The flexible β-karyopherin fold of Tnpo3 embraces the RNA recognition motif and RS domains of the cargo. A constellation of charged residues on and around the arginine-rich helix of Tnpo3 HEAT repeat 15 engage the phosphorylated RS domain and are critical for the recognition and nuclear import of ASF/SF2. Mutations in the same region of Tnpo3 impair its interaction with the cleavage and polyadenylation specificity factor 6 (CPSF6) and its ability to support HIV-1 replication. Steric incompatibility of the RS domain and RanGTP engagement by Tnpo3 provides the mechanism for cargo release in the nucleus. Our results elucidate the structural bases for nuclear import of splicing factors and the Tnpo3-CPSF6 nexus in HIV-1 biology.T he transport of macromolecules between cytoplasm and nucleus is orchestrated by a family of nuclear import and export receptors (1). Referred to as importins and exportins, these proteins bind their specific cargoes and translocate them across the nuclear pore complex. The process is regulated by the small GTPase Ran that partitions between cytoplasm and nucleus in the predominantly GDP-and GTP-bound form, respectively. Importins associate with their cargoes in the cytoplasm, and the competitive binding of RanGTP induces them to release their cargoes in the nucleus (2). Most nuclear import/export receptors belong to the β-karyopherin family of proteins, with 22 members encoded in the human genome (3). The majority of β-karyopherins bind their cargoes directly, recognizing a linear nuclear localization or export signal and/or a specific tertiary/quaternary structural feature (4, 5).A fundamentally important type of a nuclear localization signal (NLS), comprising sequences rich in Arg-Ser and/or ArgAsp/Glu/Gly dipeptides (referred to as RS or RS-like domains), belongs to the family of Ser/Arg-rich (SR) proteins. These nuclear proteins also contain RNA recognition motif (RRM) domains and play essential roles in pre-mRNA splicing and 3′ processing and participate in transcription regulation, mRNA transport, translation, and nonsense-mediated mRNA decay (6). The splicing factors alternative splicing factor/splicing factor 2 (ASF/SF2) and SC35 along with the cleavage and polyadenylation specificity factor 6 (CPSF6, also known as CF-Im-68) are among the best-characterized metazoan SR proteins (7-10). The SR protein family can be further extended by inclusion of a structurally and functionally diverse group of nuclear proteins that possess RS repeats but lack RRM domains (11). The RS domains are processively phosphorylated on their Ser residues by a set of dedicated kinases (12-16). Phosphorylation of SR proteins is thought to...

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Section: Resultssupporting

confidence: 48%

Structural basis for nuclear import of splicing factors by human Transportin 3

Maertens

Cook

Wang

et al. 2014

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

130

167

View full text Add to dashboard Cite

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“…We previously characterized the TRN-SR2 interaction interface to identify key amino acids (27). Independent confirmation of the role of Arg 262 , Arg 263 , and Lys 264 was provided by Larue et al (38).…”

Section: Discussionmentioning

confidence: 98%

“…Although we and others have shown that TRN-SR2 directly interacts with HIV IN (18,19,27,38,39), the mechanism of action has been questioned (19,23,36,40,41). Especially reports on a set of HIV capsid mutations (e.g.…”

mentioning

confidence: 99%

“…To uncouple the effect of TRN-SR2 knockdown from the possible cytoplasmic accumulation of CPSF6 or other cargoes, we decided to search for IN mutants specifically defective for TRN-SR2 binding. We previously characterized the TRN-SR2 interaction interface to identify key amino acids in the viral IN and determined the hot spots for the interaction, namely the amino acids Arg (38).…”

mentioning

confidence: 99%

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The HIV-1 Integrase Mutant R263A/K264A Is 2-fold Defective for TRN-SR2 Binding and Viral Nuclear Import

Houwer

Demeulemeester

Thys

et al. 2014

Journal of Biological Chemistry

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Background: Whether the TRN-SR2/HIV-1 IN interaction mediates the nuclear import of HIV is controversial. Results: The replication-defective HIV integrase mutant INR263A/K264A displays a 2-fold reduction in interaction with TRN-SR2 and PIC nuclear import. Conclusion:The HIV-IN TRN-SR2 protein-protein interaction is important for HIV nuclear import. Significance: The IN/TRN-SR2 interaction interface is a potential target for future antiviral therapy.

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“…Proteins and the Inhibitor-Full-length recombinant WT and mutant (A128T and K264A/K266A) INs containing the N-terminal His 6 tag were expressed in Escherichia coli and purified as described previously (49). Similar purification protocols were used to prepare isolated IN domains (CCD, CTD, and the CCD-CTD).…”

Section: Methodsmentioning

confidence: 99%

A Critical Role of the C-terminal Segment for Allosteric Inhibitor-induced Aberrant Multimerization of HIV-1 Integrase

Shkriabai

Dharmarajan

Slaughter

et al. 2014

Journal of Biological Chemistry

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Background: Allosteric integrase (IN) inhibitors (ALLINIs) promote aberrant protein multimerization. Results: ALLINI-2 induces protein-protein interactions, including C-terminal residues Lys-264 and Lys-266, which lead to aberrant, higher-order IN multimerization. Conclusion: The protein-protein contacts beyond the inhibitor binding site contribute to aberrant IN multimerization. Significance: Our findings provide structural clues and underscore the significance for exploiting IN multimerization as a new therapeutic target.

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Interaction of the HIV-1 Intasome with Transportin 3 Protein (TNPO3 or TRN-SR2)

Cited by 52 publications

References 83 publications

Structural basis for nuclear import of splicing factors by human Transportin 3

Structural basis for nuclear import of splicing factors by human Transportin 3

The HIV-1 Integrase Mutant R263A/K264A Is 2-fold Defective for TRN-SR2 Binding and Viral Nuclear Import

A Critical Role of the C-terminal Segment for Allosteric Inhibitor-induced Aberrant Multimerization of HIV-1 Integrase

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