Abstract
Nuclear import and export of viral nucleic acids is crucial for the replication cycle of many viruses, and elucidation of the mechanism of these steps may provide a paradigm for understanding general biological processes. Influenza virus replicates its RNA genome in the nucleus of infected cells. The influenza virus NS2 protein, which had no previously assigned function, was shown to mediate the nuclear export of virion RNAs by acting as an adaptor between viral ribonucleoprotein complexes and the nuclear export machinery of the cell. A functional domain on the NS2 with characteristics of a nuclear export signal was mapped: it interacts with cellular nucleoporins, can functionally replace the effector domain of the human immunodeficiency virus type 1 (HIV-1) Rev protein and mediates rapid nuclear export when cross-linked to a reporter protein. Microinjection of anti-NS2 antibodies into infected cells inhibited nuclear export of viral ribonucleoproteins, suggesting that the Rev-like NS2 mediates this process. Therefore, we have renamed this Rev-like factor the influenza virus nuclear export protein or NEP. We propose a model by which NEP acts as a protein adaptor molecule bridging viral ribonucleoproteins and the nuclear pore complex.
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- Alonso-Caplen F. V., Krug R. M. Regulation of the extent of splicing of influenza virus NS1 mRNA: role of the rates of splicing and of the nucleocytoplasmic transport of NS1 mRNA. Mol Cell Biol. 1991 Feb;11(2):1092–1098. doi: 10.1128/mcb.11.2.1092. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Alonso-Caplen F. V., Nemeroff M. E., Qiu Y., Krug R. M. Nucleocytoplasmic transport: the influenza virus NS1 protein regulates the transport of spliced NS2 mRNA and its precursor NS1 mRNA. Genes Dev. 1992 Feb;6(2):255–267. doi: 10.1101/gad.6.2.255. [DOI] [PubMed] [Google Scholar]
- Bogerd H. P., Fridell R. A., Madore S., Cullen B. R. Identification of a novel cellular cofactor for the Rev/Rex class of retroviral regulatory proteins. Cell. 1995 Aug 11;82(3):485–494. doi: 10.1016/0092-8674(95)90437-9. [DOI] [PubMed] [Google Scholar]
- Davis L. I. The nuclear pore complex. Annu Rev Biochem. 1995;64:865–896. doi: 10.1146/annurev.bi.64.070195.004245. [DOI] [PubMed] [Google Scholar]
- Diaz J. J., Dodon M. D., Schaerer-Uthurralt N., Simonin D., Kindbeiter K., Gazzolo L., Madjar J. J. Post-transcriptional transactivation of human retroviral envelope glycoprotein expression by herpes simplex virus Us11 protein. Nature. 1996 Jan 18;379(6562):273–277. doi: 10.1038/379273a0. [DOI] [PubMed] [Google Scholar]
- Dobbelstein M., Roth J., Kimberly W. T., Levine A. J., Shenk T. Nuclear export of the E1B 55-kDa and E4 34-kDa adenoviral oncoproteins mediated by a rev-like signal sequence. EMBO J. 1997 Jul 16;16(14):4276–4284. doi: 10.1093/emboj/16.14.4276. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Felber B. K., Hadzopoulou-Cladaras M., Cladaras C., Copeland T., Pavlakis G. N. rev protein of human immunodeficiency virus type 1 affects the stability and transport of the viral mRNA. Proc Natl Acad Sci U S A. 1989 Mar;86(5):1495–1499. doi: 10.1073/pnas.86.5.1495. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Fischer U., Huber J., Boelens W. C., Mattaj I. W., Lührmann R. The HIV-1 Rev activation domain is a nuclear export signal that accesses an export pathway used by specific cellular RNAs. Cell. 1995 Aug 11;82(3):475–483. doi: 10.1016/0092-8674(95)90436-0. [DOI] [PubMed] [Google Scholar]
- Fischer U., Meyer S., Teufel M., Heckel C., Lührmann R., Rautmann G. Evidence that HIV-1 Rev directly promotes the nuclear export of unspliced RNA. EMBO J. 1994 Sep 1;13(17):4105–4112. doi: 10.1002/j.1460-2075.1994.tb06728.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Fortes P., Beloso A., Ortín J. Influenza virus NS1 protein inhibits pre-mRNA splicing and blocks mRNA nucleocytoplasmic transport. EMBO J. 1994 Feb 1;13(3):704–712. doi: 10.1002/j.1460-2075.1994.tb06310.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Fridell R. A., Bogerd H. P., Cullen B. R. Nuclear export of late HIV-1 mRNAs occurs via a cellular protein export pathway. Proc Natl Acad Sci U S A. 1996 Apr 30;93(9):4421–4424. doi: 10.1073/pnas.93.9.4421. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Fridell R. A., Fischer U., Lührmann R., Meyer B. E., Meinkoth J. L., Malim M. H., Cullen B. R. Amphibian transcription factor IIIA proteins contain a sequence element functionally equivalent to the nuclear export signal of human immunodeficiency virus type 1 Rev. Proc Natl Acad Sci U S A. 1996 Apr 2;93(7):2936–2940. doi: 10.1073/pnas.93.7.2936. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Fritz C. C., Zapp M. L., Green M. R. A human nucleoporin-like protein that specifically interacts with HIV Rev. Nature. 1995 Aug 10;376(6540):530–533. doi: 10.1038/376530a0. [DOI] [PubMed] [Google Scholar]
- Greenspan D., Krystal M., Nakada S., Arnheiter H., Lyles D. S., Palese P. Expression of influenza virus NS2 nonstructural protein in bacteria and localization of NS2 in infected eucaryotic cells. J Virol. 1985 Jun;54(3):833–843. doi: 10.1128/jvi.54.3.833-843.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Guddat U., Bakken A. H., Pieler T. Protein-mediated nuclear export of RNA: 5S rRNA containing small RNPs in xenopus oocytes. Cell. 1990 Feb 23;60(4):619–628. doi: 10.1016/0092-8674(90)90665-2. [DOI] [PubMed] [Google Scholar]
- Guiochon-Mantel A., Lescop P., Christin-Maitre S., Loosfelt H., Perrot-Applanat M., Milgrom E. Nucleocytoplasmic shuttling of the progesterone receptor. EMBO J. 1991 Dec;10(12):3851–3859. doi: 10.1002/j.1460-2075.1991.tb04954.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Gyuris J., Golemis E., Chertkov H., Brent R. Cdi1, a human G1 and S phase protein phosphatase that associates with Cdk2. Cell. 1993 Nov 19;75(4):791–803. doi: 10.1016/0092-8674(93)90498-f. [DOI] [PubMed] [Google Scholar]
- Hope T. J., Huang X. J., McDonald D., Parslow T. G. Steroid-receptor fusion of the human immunodeficiency virus type 1 Rev transactivator: mapping cryptic functions of the arginine-rich motif. Proc Natl Acad Sci U S A. 1990 Oct;87(19):7787–7791. doi: 10.1073/pnas.87.19.7787. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Izaurralde E., Mattaj I. W. RNA export. Cell. 1995 Apr 21;81(2):153–159. doi: 10.1016/0092-8674(95)90323-2. [DOI] [PubMed] [Google Scholar]
- Lamb R. A., Etkind P. R., Choppin P. W. Evidence for a ninth influenza viral polypeptide. Virology. 1978 Nov;91(1):60–78. doi: 10.1016/0042-6822(78)90355-0. [DOI] [PubMed] [Google Scholar]
- Malim M. H., Böhnlein S., Hauber J., Cullen B. R. Functional dissection of the HIV-1 Rev trans-activator--derivation of a trans-dominant repressor of Rev function. Cell. 1989 Jul 14;58(1):205–214. doi: 10.1016/0092-8674(89)90416-9. [DOI] [PubMed] [Google Scholar]
- Malim M. H., Hauber J., Le S. Y., Maizel J. V., Cullen B. R. The HIV-1 rev trans-activator acts through a structured target sequence to activate nuclear export of unspliced viral mRNA. Nature. 1989 Mar 16;338(6212):254–257. doi: 10.1038/338254a0. [DOI] [PubMed] [Google Scholar]
- Malim M. H., McCarn D. F., Tiley L. S., Cullen B. R. Mutational definition of the human immunodeficiency virus type 1 Rev activation domain. J Virol. 1991 Aug;65(8):4248–4254. doi: 10.1128/jvi.65.8.4248-4254.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Martin K., Helenius A. Nuclear transport of influenza virus ribonucleoproteins: the viral matrix protein (M1) promotes export and inhibits import. Cell. 1991 Oct 4;67(1):117–130. doi: 10.1016/0092-8674(91)90576-k. [DOI] [PubMed] [Google Scholar]
- Mehlin H., Daneholt B., Skoglund U. Translocation of a specific premessenger ribonucleoprotein particle through the nuclear pore studied with electron microscope tomography. Cell. 1992 May 15;69(4):605–613. doi: 10.1016/0092-8674(92)90224-z. [DOI] [PubMed] [Google Scholar]
- Meyer B. E., Meinkoth J. L., Malim M. H. Nuclear transport of human immunodeficiency virus type 1, visna virus, and equine infectious anemia virus Rev proteins: identification of a family of transferable nuclear export signals. J Virol. 1996 Apr;70(4):2350–2359. doi: 10.1128/jvi.70.4.2350-2359.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Michael W. M., Choi M., Dreyfuss G. A nuclear export signal in hnRNP A1: a signal-mediated, temperature-dependent nuclear protein export pathway. Cell. 1995 Nov 3;83(3):415–422. doi: 10.1016/0092-8674(95)90119-1. [DOI] [PubMed] [Google Scholar]
- Murphy R., Wente S. R. An RNA-export mediator with an essential nuclear export signal. Nature. 1996 Sep 26;383(6598):357–360. doi: 10.1038/383357a0. [DOI] [PubMed] [Google Scholar]
- Nigg E. A. Nucleocytoplasmic transport: signals, mechanisms and regulation. Nature. 1997 Apr 24;386(6627):779–787. doi: 10.1038/386779a0. [DOI] [PubMed] [Google Scholar]
- O'Neill R. E., Jaskunas R., Blobel G., Palese P., Moroianu J. Nuclear import of influenza virus RNA can be mediated by viral nucleoprotein and transport factors required for protein import. J Biol Chem. 1995 Sep 29;270(39):22701–22704. doi: 10.1074/jbc.270.39.22701. [DOI] [PubMed] [Google Scholar]
- O'Neill R. E., Palese P. NPI-1, the human homolog of SRP-1, interacts with influenza virus nucleoprotein. Virology. 1995 Jan 10;206(1):116–125. doi: 10.1016/s0042-6822(95)80026-3. [DOI] [PubMed] [Google Scholar]
- Palmeri D., Malim M. H. The human T-cell leukemia virus type 1 posttranscriptional trans-activator Rex contains a nuclear export signal. J Virol. 1996 Sep;70(9):6442–6445. doi: 10.1128/jvi.70.9.6442-6445.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Piñol-Roma S., Dreyfuss G. Shuttling of pre-mRNA binding proteins between nucleus and cytoplasm. Nature. 1992 Feb 20;355(6362):730–732. doi: 10.1038/355730a0. [DOI] [PubMed] [Google Scholar]
- Qiu Y., Krug R. M. The influenza virus NS1 protein is a poly(A)-binding protein that inhibits nuclear export of mRNAs containing poly(A). J Virol. 1994 Apr;68(4):2425–2432. doi: 10.1128/jvi.68.4.2425-2432.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Rey O., Nayak D. P. Nuclear retention of M1 protein in a temperature-sensitive mutant of influenza (A/WSN/33) virus does not affect nuclear export of viral ribonucleoproteins. J Virol. 1992 Oct;66(10):5815–5824. doi: 10.1128/jvi.66.10.5815-5824.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Richardson J. C., Akkina R. K. NS2 protein of influenza virus is found in purified virus and phosphorylated in infected cells. Arch Virol. 1991;116(1-4):69–80. doi: 10.1007/BF01319232. [DOI] [PubMed] [Google Scholar]
- Stutz F., Izaurralde E., Mattaj I. W., Rosbash M. A role for nucleoporin FG repeat domains in export of human immunodeficiency virus type 1 Rev protein and RNA from the nucleus. Mol Cell Biol. 1996 Dec;16(12):7144–7150. doi: 10.1128/mcb.16.12.7144. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Stutz F., Neville M., Rosbash M. Identification of a novel nuclear pore-associated protein as a functional target of the HIV-1 Rev protein in yeast. Cell. 1995 Aug 11;82(3):495–506. doi: 10.1016/0092-8674(95)90438-7. [DOI] [PubMed] [Google Scholar]
- Stutz F., Rosbash M. A functional interaction between Rev and yeast pre-mRNA is related to splicing complex formation. EMBO J. 1994 Sep 1;13(17):4096–4104. doi: 10.1002/j.1460-2075.1994.tb06727.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Visa N., Alzhanova-Ericsson A. T., Sun X., Kiseleva E., Björkroth B., Wurtz T., Daneholt B. A pre-mRNA-binding protein accompanies the RNA from the gene through the nuclear pores and into polysomes. Cell. 1996 Jan 26;84(2):253–264. doi: 10.1016/s0092-8674(00)80980-0. [DOI] [PubMed] [Google Scholar]
- Ward A. C., Castelli L. A., Lucantoni A. C., White J. F., Azad A. A., Macreadie I. G. Expression and analysis of the NS2 protein of influenza A virus. Arch Virol. 1995;140(11):2067–2073. doi: 10.1007/BF01322693. [DOI] [PubMed] [Google Scholar]
- Wen W., Harootunian A. T., Adams S. R., Feramisco J., Tsien R. Y., Meinkoth J. L., Taylor S. S. Heat-stable inhibitors of cAMP-dependent protein kinase carry a nuclear export signal. J Biol Chem. 1994 Dec 23;269(51):32214–32220. [PubMed] [Google Scholar]
- Wen W., Meinkoth J. L., Tsien R. Y., Taylor S. S. Identification of a signal for rapid export of proteins from the nucleus. Cell. 1995 Aug 11;82(3):463–473. doi: 10.1016/0092-8674(95)90435-2. [DOI] [PubMed] [Google Scholar]
- Whittaker G., Bui M., Helenius A. Nuclear trafficking of influenza virus ribonuleoproteins in heterokaryons. J Virol. 1996 May;70(5):2743–2756. doi: 10.1128/jvi.70.5.2743-2756.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Whittaker G., Bui M., Helenius A. The role of nuclear import and export in influenza virus infection. Trends Cell Biol. 1996 Feb;6(2):67–71. doi: 10.1016/0962-8924(96)81017-8. [DOI] [PubMed] [Google Scholar]
- Wolff T., O'Neill R. E., Palese P. Interaction cloning of NS1-I, a human protein that binds to the nonstructural NS1 proteins of influenza A and B viruses. J Virol. 1996 Aug;70(8):5363–5372. doi: 10.1128/jvi.70.8.5363-5372.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Yasuda J., Nakada S., Kato A., Toyoda T., Ishihama A. Molecular assembly of influenza virus: association of the NS2 protein with virion matrix. Virology. 1993 Sep;196(1):249–255. doi: 10.1006/viro.1993.1473. [DOI] [PubMed] [Google Scholar]
- Zervos A. S., Gyuris J., Brent R. Mxi1, a protein that specifically interacts with Max to bind Myc-Max recognition sites. Cell. 1993 Jan 29;72(2):223–232. doi: 10.1016/0092-8674(93)90662-a. [DOI] [PubMed] [Google Scholar]