trans-Activation of the HIV-1 LTR by the HIV-1 tat and HTLV-I tax proteins is mediated by different cis-acting sequences

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.

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HIV- 1: To Splice Or Not To Splice, That Is THE Question

10.20944/preprints202101.0114.v1 ◽

2021 ◽

Author(s):

Ronald Swanstrom ◽

Ann Emery

Keyword(s):

Fitness Cost ◽

Full Length ◽

Regulatory Sequences ◽

Supporting Evidence ◽

Control Mechanisms ◽

Genomic Rna ◽

Cis Acting ◽

Splicing Regulators ◽

Mrna Transcripts ◽

Hiv 1

HIV-1 transcribes only one kind of transcript – the full length genomic RNA. To make the mRNA transcripts for the accessory proteins Tat and Rev, the genomic RNA must completely splice. The mRNA transcripts for Vif, Vpr, and Env must splice but not completely. Genomic RNA (which also functions as mRNA for the Gag and Gag/Pro/Pol precursor polyproteins) must not splice at all. HIV-1 can tolerate a surprising range in the relative abundance of individual transcript types, and a surprising amount of aberrant and even odd splicing; however, it must not over-splice, which results in the loss of full length genomic RNA and has a dramatic fitness cost. Loss of full length genomic transcripts through over-splicing has a dramatic fitness cost. Cells typically do not tolerate unspliced/incompletely spliced transcripts, so HIV-1 has to circumvent this cell policing mechanism to allow some splicing while suppressing most. Splicing is controlled by RNA secondary structure, cis-acting regulatory sequences which bind splicing factors, and the viral protein Rev. There is still much work to be done to clarify the combinatorial effects of these splicing regulators. These control mechanisms represent attractive targets to induce over-splicing as an antiviral strategy. Finally, splicing has been implicated in latency, but to date there is little supporting evidence for such a mechanism. In this review we apply what is known of cellular splicing to understand splicing in HIV-1, and also present data from our newer and more sensitive deep sequencing assays quantifying the different HIV-1 transcript types.

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Importance of independence in ribozyme reactions: kinetic behavior of trimmed and of simply connected multiple ribozymes with potential activity against human immunodeficiency virus

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.90.23.11302 ◽

1993 ◽

Vol 90 (23) ◽

pp. 11302-11306 ◽

Cited By ~ 51

Author(s):

J Ohkawa ◽

N Yuyama ◽

Y Takebe ◽

S Nishikawa ◽

K Taira

Keyword(s):

Human Immunodeficiency Virus ◽

Expression Vector ◽

Expression System ◽

Kinetic Behavior ◽

Cis Acting ◽

Immunodeficiency Virus ◽

Potential Activity ◽

Simply Connected ◽

Hiv 1

The kinetic behavior of ribozymes derived from two types of multiple-ribozyme expression vector were examined. In some cases, multiple ribozymes were expressed as a single RNA molecule and all the ribozymes were simply connected in tandem (connected type). In other cases, multiple ribozymes were flanked by cis-acting ribozymes at both their 5' and 3' ends so that, upon transcription, multiple ribozymes were trimmed at both their 5' and 3' ends, with resultant liberation of multiple independent ribozymes (shotgun type). When levels of ribozyme expression were examined for the shotgun-type vector, the level of the ribozyme transcript was found to be proportional to the number of units (n) connected in tandem. Accordingly, the activities of the shotgun-type ribozymes, in terms of the cleavage of HIV-1 RNA in vitro, were also found to be proportional to the number of units connected in tandem (n). By contrast, the activities of the connected-type ribozymes reached plateau values at around n = 3. These results indicate that, when the shotgun-type expression system is used, it is theoretically possible to generate various independent ribozymes, each specific for a different target site, without sacrificing the activity of any individual ribozyme.

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HIV-1: To Splice or Not to Splice, That Is the Question

Viruses ◽

10.3390/v13020181 ◽

2021 ◽

Vol 13 (2) ◽

pp. 181

Author(s):

Ann Emery ◽

Ronald Swanstrom

Keyword(s):

Full Length ◽

Regulatory Sequences ◽

Supporting Evidence ◽

Control Mechanisms ◽

Genomic Rna ◽

Cis Acting ◽

Splicing Regulators ◽

Individual Transcript ◽

Mrna Transcripts ◽

Hiv 1

The transcription of the HIV-1 provirus results in only one type of transcript—full length genomic RNA. To make the mRNA transcripts for the accessory proteins Tat and Rev, the genomic RNA must completely splice. The mRNA transcripts for Vif, Vpr, and Env must undergo splicing but not completely. Genomic RNA (which also functions as mRNA for the Gag and Gag/Pro/Pol precursor polyproteins) must not splice at all. HIV-1 can tolerate a surprising range in the relative abundance of individual transcript types, and a surprising amount of aberrant and even odd splicing; however, it must not over-splice, which results in the loss of full-length genomic RNA and has a dramatic fitness cost. Cells typically do not tolerate unspliced/incompletely spliced transcripts, so HIV-1 must circumvent this cell policing mechanism to allow some splicing while suppressing most. Splicing is controlled by RNA secondary structure, cis-acting regulatory sequences which bind splicing factors, and the viral protein Rev. There is still much work to be done to clarify the combinatorial effects of these splicing regulators. These control mechanisms represent attractive targets to induce over-splicing as an antiviral strategy. Finally, splicing has been implicated in latency, but to date there is little supporting evidence for such a mechanism. In this review we apply what is known of cellular splicing to understand splicing in HIV-1, and present data from our newer and more sensitive deep sequencing assays quantifying the different HIV-1 transcript types.

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A multifunctional expression vector for an anti-HIV-1 ribozyme that produces a 5′- and 3′-trimmed trans-acting ribozyme, targeted against HIV-1 RNA, and cis-acting ribozymes that are designed to bind to and thereby sequester frans-activator proteins such as Tat and Rev

Nucleic Acids Research ◽

10.1093/nar/22.23.5060 ◽

1994 ◽

Vol 22 (23) ◽

pp. 5060-5067 ◽

Cited By ~ 13

Author(s):

Noriko Yuyama ◽

Jun Ohkawa ◽

Tetsuhiko Koguma ◽

Makoto Shirai ◽

Kazunari Taira

Keyword(s):

Expression Vector ◽

Cis Acting ◽

Activator Proteins ◽

Anti Hiv ◽

Hiv 1

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Let it go: HIV-1 cis-acting repressive sequences

Journal of Virology ◽

10.1128/jvi.00342-21 ◽

2021 ◽

Author(s):

Philipp Niklas Ostermann ◽

Anastasia Ritchie ◽

Johannes Ptok ◽

Heiner Schaal

Keyword(s):

Nuclear Export ◽

Negative Impact ◽

Cis Acting ◽

Bioinformatic Tools ◽

Cellular Genes ◽

Immunodeficiency Virus ◽

Underlying Mechanisms ◽

Predictive Approaches ◽

Intensive Work ◽

Hiv 1

After human immunodeficiency virus type 1 (HIV-1) was identified in the early 1980s, intensive work began to understand the molecular basis of HIV-1 gene expression. Subgenomic HIV-1 RNA regions, spread throughout the viral genome, were described to have a negative impact on the nuclear export of some viral transcripts. These studies revealed an intrinsic RNA code as a new form of nuclear export regulation. Since such regulatory regions were later also identified in other viruses as well as in cellular genes, it can be assumed that during evolution, viruses took advantage of them to achieve more sophisticated replication mechanisms. Here, we review HIV-1 cis-acting repressive sequences that have been identified and discuss their possible underlying mechanisms and importance. Additionally, we show how current bioinformatic tools might allow more predictive approaches to identify and investigate them.

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HIV-1 Vpr binding to HIV-1 LTR C/EBP cis-acting elements and adjacent regions is sequence-specific

Biomedicine & Pharmacotherapy ◽

10.1016/s0753-3322(02)00333-5 ◽

2003 ◽

Vol 57 (1) ◽

pp. 41-48 ◽

Cited By ~ 22

Author(s):

Tricia H. Hogan ◽

Michael R. Nonnemacher ◽

Fred C. Krebs ◽

Andrew Henderson ◽

Brian Wigdahl

Keyword(s):

Cis Acting ◽

Hiv 1

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Presence of negative and positive cis-acting RNA splicing elements within and flanking the first tat coding exon of human immunodeficiency virus type 1.

Molecular and Cellular Biology ◽

10.1128/mcb.14.6.3960 ◽

1994 ◽

Vol 14 (6) ◽

pp. 3960-3970 ◽

Cited By ~ 80

Author(s):

B A Amendt ◽

D Hesslein ◽

L J Chang ◽

C M Stoltzfus

Keyword(s):

Human Immunodeficiency Virus ◽

Human Immunodeficiency Virus Type ◽

Virus Type ◽

Splice Site ◽

Rna Splicing ◽

Regulatory Element ◽

Cis Acting ◽

Immunodeficiency Virus ◽

Hiv 1

The human immunodeficiency virus type 1 (HIV-1) RNA follows a complex splicing pathway in which a single primary transcript either remains unspliced or is alternatively spliced to more than 30 different singly and multiply spliced mRNAs. We have used an in vitro splicing assay to identify cis elements within the viral genome that regulate HIV-1 RNA splicing. A novel splicing regulatory element (SRE) within the first tat coding exon has been detected. This element specifically inhibits splicing at the upstream 3' splice site flanking this tat exon. The element only functions when in the sense orientation and is position dependent when inserted downstream of a heterologous 3' splice site. In vivo, an HIV-1 SRE mutant demonstrated a decrease in unspliced viral RNA, increased levels of single- and double-spliced tat mRNA, and reduced levels of env and rev mRNAs. In addition to the negative cis-acting SRE, the flanking 5' splice site downstream of the first tat coding exon acts positively to increase splicing at the upstream 3' splice sites. These results are consistent with hypotheses of bridging interactions between cellular factors that bind to the 5' splice site and those that bind at the upstream 3' splice site.

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Identification of a Novel Cis-Acting Regulator of HIV-1 Genome Packaging

International Journal of Molecular Sciences ◽

10.3390/ijms22073435 ◽

2021 ◽

Vol 22 (7) ◽

pp. 3435

Author(s):

Sayuri Sakuragi ◽

Osamu Kotani ◽

Masaru Yokoyama ◽

Tatsuo Shioda ◽

Hironori Sato ◽

...

Keyword(s):

Structural Changes ◽

Viral Rna ◽

Psi Function ◽

Genome Packaging ◽

Cis Acting ◽

Binding Interface ◽

Subtype B ◽

Hydrogen Bond Networks ◽

Genome Dimerization ◽

Hiv 1

Human immunodeficiency virus type 1 (HIV-1) uptakes homo-dimerized viral RNA genome into its own particle. A cis-acting viral RNA segment responsible for this event, termed packaging signal (psi), is located at the 5′-end of the viral genome. Although the psi segment exhibits nucleotide variation in nature, its effects on the psi function largely remain unknown. Here we show that a psi sequence from an HIV-1 regional variant, subtype D, has a lower packaging ability compared with that from another regional variant, HIV-1 subtype B, despite maintaining similar genome dimerization activities. A series of molecular genetic investigations narrowed down the responsible element of the selective attenuation to the two sequential nucleotides at positions 226 and 227 in the psi segment. Molecular dynamics simulations predicted that the dinucleotide substitution alters structural dynamics, fold, and hydrogen-bond networks primarily of the psi-SL2 element that contains the binding interface of viral nucleocapsid protein for the genome packaging. In contrast, such structural changes were minimal within the SL1 element involved in genome dimerization. These results suggest that the psi 226/227 dinucleotide pair functions as a cis-acting regulator to control the psi structure to selectively tune the efficiency of packaging, but not dimerization of highly variable HIV-1 genomes.

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