scholarly journals Concerted Action of Multiple cis-Acting Sequences Is Required for Rev Dependence of Late Human Immunodeficiency Virus Type 1 Gene Expression

2000 ◽  
Vol 74 (22) ◽  
pp. 10822-10826 ◽  
Author(s):  
Marcus Graf ◽  
Alexandra Bojak ◽  
Ludwig Deml ◽  
Kurt Bieler ◽  
Hans Wolf ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Nuclear Export ◽  
Regulatory Elements ◽  
Cis Acting ◽  
Immunodeficiency Virus ◽  
Responsive Element ◽  
Hiv 1

ABSTRACT Based on the human immunodeficiency virus type 1 (HIV-1)gag gene, subgenomic reporter constructs have been established allowing the contributions of differentcis-acting elements to the Rev dependency of late HIV-1 gene products to be determined. Modification of intragenic regulatory elements achieved by adapting the codon usage of the complete gene to highly expressed mammalian genes resulted in constitutive nuclear export allowing high levels of Gag expression independent from the Rev/Rev-responsive element system and irrespective of the absence or presence of the isolated major splice donor. Leptomycin B inhibitor studies revealed that the RNAs derived from the codon-optimizedgag gene lacking AU-rich inhibitory elements are directed to a distinct, CRM1-independent, nuclear export pathway.

scholarly journals PSF Acts through the Human Immunodeficiency Virus Type 1 mRNA Instability Elements To Regulate Virus Expression

2003 ◽  
Vol 23 (18) ◽  
pp. 6618-6630 ◽  
Author(s):  
Andrei S. Zolotukhin ◽  
Daniel Michalowski ◽  
Jenifer Bear ◽  
Sergey V. Smulevitch ◽  
Abdulmaged M. Traish ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Nuclear Export ◽  
Regulatory Elements ◽  
Nucleocytoplasmic Transport ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Human immunodeficiency virus type 1 (HIV) gag/pol and env mRNAs contain cis-acting regulatory elements (INS) that impair stability, nucleocytoplasmic transport, and translation by unknown mechanisms. This downregulation can be counteracted by the viral Rev protein, resulting in efficient export and expression of these mRNAs. Here, we show that the INS region in HIV-1 gag mRNA is a high-affinity ligand of p54nrb/PSF, a heterodimeric transcription/splicing factor. Both subunits bound INS RNA in vitro with similar affinity and specificity. Using an INS-containing subgenomic gag mRNA, we show that it specifically associated with p54nrb in vivo and that PSF inhibited its expression, acting via INS. Studying the authentic HIV-1 mRNAs produced from an infectious molecular clone, we found that PSF affected specifically the INS-containing, Rev-dependent transcripts encoding Gag-Pol and Env. Both subunits contained nuclear export and nuclear retention signals, whereas p54nrb was continuously exported from the nucleus and associated with INS-containing mRNA in the cytoplasm, suggesting its additional role at late steps of mRNA metabolism. Thus, p54nrb and PSF have properties of key factors mediating INS function and likely define a novel mRNA regulatory pathway that is hijacked by HIV-1.

scholarly journals Negative and Positive mRNA Splicing Elements Act Competitively To Regulate Human Immunodeficiency Virus Type 1 Vif Gene Expression

2008 ◽  
Vol 82 (8) ◽  
pp. 3921-3931 ◽  
Author(s):  
C. M. Exline ◽  
Z. Feng ◽  
C. M. Stoltzfus
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Regulatory Elements ◽  
Mrna Splicing ◽  
Viral Mrnas ◽  
Exon 2 ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Over 40 different human immunodeficiency virus type 1 (HIV-1) mRNAs are produced by alternative splicing of the primary HIV-1 RNA transcripts. In addition, approximately half of the viral RNA remains unspliced and is used as genomic RNA and as mRNA for the Gag and Pol gene products. Regulation of splicing at the HIV-1 3′ splice sites (3′ss) requires suboptimal polypyrimidine tracts, and positive or negative regulation occurs through the binding of cellular factors to cis-acting splicing regulatory elements. We have previously shown that splicing at HIV-1 3′ss A1, which produces single-spliced vif mRNA and promotes the inclusion of HIV exon 2 into both completely and incompletely spliced viral mRNAs, is increased by optimizing the 5′ splice site (5′ss) downstream of exon 2 (5′ss D2). Here we show that the mutations within 5′ss D2 that are predicted to lower or increase the affinity of the 5′ss for U1 snRNP result in reduced or increased Vif expression, respectively. Splicing at 5′ss D2 was not necessary for the effect of 5′ss D2 on Vif expression. In addition, we have found that mutations of the GGGG motif proximal to the 5′ss D2 increase exon 2 inclusion and Vif expression. Finally, we report the presence of a novel exonic splicing enhancer (ESE) element within the 5′-proximal region of exon 2 that facilitates both exon inclusion and Vif expression. This ESE binds specifically to the cellular SR protein SRp75. Our results suggest that the 5′ss D2, the proximal GGGG silencer, and the ESE act competitively to determine the level of vif mRNA splicing and Vif expression. We propose that these positive and negative splicing elements act together to allow the accumulation of vif mRNA and unspliced HIV-1 mRNA, compatible with optimal virus replication.

scholarly journals SRp40 and SRp55 Promote the Translation of Unspliced Human Immunodeficiency Virus Type 1 RNA

2010 ◽  
Vol 84 (13) ◽  
pp. 6748-6759 ◽  
Author(s):  
Chad M. Swanson ◽  
Nathan M. Sherer ◽  
Michael H. Malim
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Nuclear Export ◽  
Sr Proteins ◽  
Rna Recognition Motif ◽  
Genomic Rnas ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Nuclear RNA processing events, such as 5′ cap formation, 3′ polyadenylation, and pre-mRNA splicing, mark mRNA for efficient translation. Splicing enhances translation via the deposition of the exon-junction complex and other multifunctional splicing factors, including SR proteins. All retroviruses synthesize their structural and enzymatic proteins from unspliced genomic RNAs (gRNAs) and must therefore exploit unconventional strategies to ensure their effective expression. Here, we report that specific SR proteins, particularly SRp40 and SRp55, promote human immunodeficiency virus type 1 (HIV-1) Gag translation from unspliced (intron-containing) viral RNA. This activity does not correlate with nucleocytoplasmic shuttling capacity and, in the case of SRp40, is dependent on the second RNA recognition motif and the arginine-serine (RS) domain. While SR proteins enhance Gag expression independent of RNA nuclear export pathway choice, altering the nucleotide sequence of the gag-pol coding region by codon optimization abolishes this effect. We therefore propose that SR proteins couple HIV-1 gRNA biogenesis to translational utilization.

scholarly journals Restriction of Human Immunodeficiency Virus Type 1 Rev Function in Murine A9 Cells Involves the Rev C-Terminal Domain

2003 ◽  
Vol 77 (5) ◽  
pp. 3084-3090 ◽  
Author(s):  
Sandra M. P. Marques ◽  
Jean-Luc Veyrune ◽  
Ram R. Shukla ◽  
Ajit Kumar
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Nuclear Export ◽  
Productive Infection ◽  
Cell Leukemia Virus Type ◽  
Terminal Domain ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT The human immunodeficiency virus type 1 (HIV-1) Rev and human T-cell leukemia virus type 1 (HTLV-1) Rex proteins are essential for the expression of viral structural proteins and productive infection. Both contain a nuclear export signal (NES) in their C-terminal domain and a nuclear localization signal (NLS) in their N-terminal domain. The NES and NLS are necessary for shuttling between nucleus and cytoplasm and are therefore indispensable for the transport of unspliced and singly spliced viral transcripts. HIV-1 Rev function is restricted in A9 cells, a murine fibroblast cell line, whereas HTLV-1 Rex is functional in these cells. Immunofluorescence studies with RevGFP fusion protein demonstrate normal import and export of Rev in A9 cells. To ascertain which domains of Rev are necessary for the restriction of Rev function in A9 cells, we studied a chimeric construct in which the NES domain of Rev was exchanged with Rex C-terminal amino acids 79 to 95, the Rev1-79/Rex79-95 chimera, which restored Rev function in A9 cells. In addition, overexpression of a truncated Rev containing the Rev C-terminal domain in the presence of wild-type Rev, led to restoration of Rev function in A9 cells. These results suggest that the C-terminal domain of HIV-1 Rev plays an important role in restricting Rev function in murine cells.

scholarly journals Inhibition of Human Immunodeficiency Virus Type 1 (HIV-1) Replication by HIV-1-Based Lentivirus Vectors Expressing Transdominant Rev

2001 ◽  
Vol 75 (8) ◽  
pp. 3590-3599 ◽  
Author(s):  
Mario R. Mautino ◽  
Nicholas Keiser ◽  
Richard A. Morgan
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Nuclear Export ◽  
Wild Type Virus ◽  
Wild Type ◽  
Helper Plasmid ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Retrovirus vectors expressing transdominant-negative mutants of Rev (TdRev) inhibit human immunodeficiency virus type 1 (HIV-1) replication by preventing the nuclear export of unspliced viral transcripts, thus inhibiting the synthesis of Gag-Pol, Env, and genomic RNA. The use of HIV-1–based vectors to express TdRev would have the advantage of allowing access to nondividing hematopoietic cells. It would also provide additional levels of protection by sequestering the viral regulatory proteins Tat and Rev, competing for encapsidation into wild-type virions, and inhibiting reverse transcription. Here we describe HIV-1-based vectors that express TdRev. These vectors contain mutations in the splicing signals or replacement of the Rev-responsive element by the simian retrovirus type 1 constitutive transport element, making them less sensitive to the inhibitory effects of TdRev. In addition, overexpression of Rev and the use of an HIV-1 helper plasmid that drives high levels of Gag-Pol synthesis were used to transiently overcome the inhibition by TdRev of the synthesis of Gag-Pol during vector production. SupT1 cells transduced with these vectors were more resistant to HIV-1 replication than cells transduced with Moloney murine leukemia virus-based vectors expressing TdRev. Furthermore, we show that these vectors can be mobilized by the wild-type virus, reducing the infectivity of virions escaping inhibition and conferring protection against HIV-1 replication to previously untransduced cells.

scholarly journals Nuclear Export of Vpr Is Required for Efficient Replication of Human Immunodeficiency Virus Type 1 in Tissue Macrophages

2003 ◽  
Vol 77 (13) ◽  
pp. 7582-7589 ◽  
Author(s):  
Michael P. Sherman ◽  
Carlos M. C. de Noronha ◽  
Lauren A. Eckstein ◽  
Jason Hataye ◽  
Pamela Mundt ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Nuclear Import ◽  
Nuclear Export ◽  
Nucleocytoplasmic Shuttling ◽  
Viral Spread ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Retroviruses must gain access to the host cell nucleus for subsequent replication and viral propagation. Human immunodeficiency virus type 1 (HIV-1) and other primate lentiviruses are distinguished from the gammaretroviruses by their ability to infect nondividing cells such as macrophages, an important viral reservoir in vivo. Rather than requiring nuclear membrane breakdown during cell division, the HIV-1 preintegration complex (PIC) enters the nucleus by traversing the central aqueous channel of the limiting nuclear pore complex. The HIV-1 PIC contains three nucleophilic proteins, matrix, integrase, and Vpr, all of which have been implicated in nuclear targeting. The mechanism by which Vpr can display such nucleophilic properties and yet also be available for incorporation into virions assembling at the plasma membrane is unresolved. We recently characterized Vpr as a nucleocytoplasmic shuttling protein that contains two novel nuclear import signals and an exportin-1-dependent nuclear export signal (NES). We now demonstrate that mutation of this NES impairs the incorporation of Vpr into newly formed virions. Furthermore, we find that the Vpr NES is required for efficient HIV replication in tissue macrophages present in human spleens and tonsils. These findings underscore how the nucleocytoplasmic shuttling of Vpr not only contributes to nuclear import of the HIV-1 PIC but also enables Vpr to be present in the cytoplasm for incorporation into virions, leading to enhancement of viral spread within nondividing tissue macrophages.

scholarly journals A Nuclear Kinesin-Like Protein Interacts with and Stimulates the Activity of the Leucine-Rich Nuclear Export Signal of the Human Immunodeficiency Virus Type 1 Rev Protein

2003 ◽  
Vol 77 (13) ◽  
pp. 7236-7243 ◽  
Author(s):  
L. K. Venkatesh ◽  
T. Gettemeier ◽  
G. Chinnadurai
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Nuclear Export ◽  
Cell Leukemia Virus Type ◽  
Rev Response Element ◽  
Immunodeficiency Virus ◽  
Hiv 1 ◽  
Rev Protein

ABSTRACT The Rev protein of human immunodeficiency virus type 1 (HIV-1) is essential for the nucleocytoplasmic transport of unspliced and partially spliced HIV mRNAs containing the Rev response element (RRE). In a yeast two-hybrid screen of a HeLa cell-derived cDNA expression library for human factors interacting with the Rev leucine-rich nuclear export sequence (NES), we identified a kinesin-like protein, REBP (Rev/Rex effector binding protein), highly homologous to Kid, the carboxy-terminal 75-residue region of which interacts specifically with the NESs of HIV-1 Rev, human T-cell leukemia virus type 1 Rex, and equine infectious anemia virus Rev but not with functionally inactive mutants thereof. REBP is a nuclear protein that colocalizes with Rev in the nucleoplasm and nuclear periphery of transfected cells. Specific, albeit weak, interaction between REBP and Rev could be demonstrated in coimmunoprecipitation assays in BSC-40 cells. REBP can modestly enhance Rev-dependent RRE-linked reporter gene expression both independently and in cooperation with the nucleoporin cofactor Rab/hRIP. Thus, REBP displays the characteristics expected of an authentic mediator of Rev NES function and may play a role in RRE RNA transport during HIV infection.

Sign in / Sign up

Export Citation Format

Share Document