The efficiency of RNA 3'-end formation is determined by the distance between the cap site and the poly(A) site in spleen necrosis virus.

Iwasaki, Katsurô; Temin, Howard M.

doi:10.1101/gad.4.12b.2299

Cited by 45 publications

(27 citation statements)

References 30 publications

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“…We suspect that splicing signals boost the levels of poly(A)+ RNA by at least two mechanisms. First, stimulation of total poly(A)+ mRNA levels in the AU3-L3 and U3-L3 series may reflect a general coupling of splicing, 3' end processing, and nuclear export (5,13,22 (15,23). How do these findings relate to the control of mRNA levels in the HIV-1 provirus?…”

Section: Discussionmentioning

confidence: 99%

Relative roles of signals upstream of AAUAAA and promoter proximity in regulation of human immunodeficiency virus type 1 mRNA 3' end formation.

1992

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Section: Discussionmentioning

confidence: 99%

Relative roles of signals upstream of AAUAAA and promoter proximity in regulation of human immunodeficiency virus type 1 mRNA 3' end formation.

1992

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“…A DNA fragment containing the SV40 polyadenylation signal was then inserted into these plasmids. It had been shown previously that the function of the SV40 polyadenylation signal was not affected by SNV R when the two sequences were present in tandem (18)(19)(20).…”

Section: Resultsmentioning

confidence: 99%

Effects of Homology Length in the Repeat Region on Minus-Strand DNA Transfer and Retroviral Replication

Dang

2001

J Virol

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Homology between the two repeat (R) regions in the retroviral genome mediates minus-strand DNA transfer during reverse transcription. We sought to define the effects of R homology lengths on minus-strand DNA transfer. We generated five murine leukemia virus (MLV)-based vectors that contained identical sequences but different lengths of the 3 R (3, 6, 12, 24 and 69 nucleotides [nt]); 69 nt is the full-length MLV R. After one round of replication, viral titers from the vector with a full-length downstream R were compared with viral titers generated from the other four vectors with reduced R lengths. Viral titers generated from vectors with R lengths reduced to one-third (24 nt) or one-sixth (12 nt) that of the wild type were not significantly affected; however, viral titers generated from vectors with only 3-or 6-nt homology in the R region were significantly lower. Because expression and packaging of the RNA were similar among all the vectors, the differences in the viral titers most likely reflected the impact of the homology lengths on the efficiency of minus-strand DNA transfer. The molecular nature of minus-strand DNA transfer was characterized in 63 proviruses. Precise R-to-R transfer was observed in most proviruses generated from vectors with 12-, 24-, or 69-nt homology in R, whereas aberrant transfers were predominantly used to generate proviruses from vectors with 3-or 6-nt homology. Reverse transcription using RNA transcribed from an upstream promoter, termed read-in RNA transcripts, resulted in most of the aberrant transfers. These data demonstrate that minus-strand DNA transfer is homology driven and a minimum homology length is required for accurate and efficient minusstrand DNA transfer.Retroviruses are RNA viruses that replicate through a DNA intermediate (49). Most retroviral particles contain viral RNA; upon infection of the target cells, viral RNA is copied into DNA by the viral enzyme reverse transcriptase (RT) (2, 27, 51) and then integrates into the target cell genome to form a provirus (49). Host cell RNA polymerase II transcribes the provirus to generate viral RNA transcripts; the full-length viral RNA is packaged into viral particles to serve as genetic material for the next round of viral infection (7).Retroviruses have evolved to adapt to the dual phase of the life cycle. Two of the adapted features are the genome structure and the mechanism by which the viral DNA is synthesized. The proviral genome contains two long terminal repeats (LTRs), one at each end of the viral DNA sequences (7, 50). The LTR is composed of three sections, unique 3Ј (U3), repeat (R), and unique 5Ј (U5) regions; each plays important roles during the viral life cycle (7). The U3 region contains the promoter from which viral RNA transcripts are expressed (7, 36). R and U5 are both important in the process of reverse transcription of the viral RNA into DNA, U5 for the initiation and R for the extension of viral DNA synthesis (7,26,48). The viral RNA transcript, which includes sequences from the upstream R to the end o...

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“…3 and quantitated in Table 1 reduction of the ATP concentration from 1 mM to 250 R±M. (22,40). For HIV-1, distances greater than 240 nucleotides from cap to AAUAAA were suggested to be necessary for efficient utilization in vivo (40).…”

Section: Resultsmentioning

confidence: 99%

Elements upstream of the AAUAAA within the human immunodeficiency virus polyadenylation signal are required for efficient polyadenylation in vitro.

1992

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Recent in vivo studies have identified specific sequences between 56 and 93 nucleotides upstream of a polyadenylation [poly(A)] consensus sequence, AAUAAA, in human immunodeficiency virus type 1 (HIV-1) that affect the efficiency of 3'-end processing at this site (A. Valsamakis, S. Zeichner, S. Carswell, and J. C. Alwine, Proc. Natl. Acad. Sci. USA 88:2108-2112, 1991). We have used HeLa cell nuclear extracts and precursor RNAs bearing the HIV-1 poly(A) signal to study the role of upstream sequences in vitro. Precursor RNAs containing the HIV-1 AAUAAA and necessary upstream (U3 region) and downstream (U5 region) sequences directed accurate cleavage and polyadenylation in vitro. The in vitro requirement for upstream sequences was demonstrated by using deletion and linker substitution mutations. The data showed that sequences between 56 and 93 nucleotides upstream of AAUAAA, which were required for efficient polyadenylation in vivo, were also required for efficient cleavage and polyadenylation in vitro. This is the first demonstration of the function of upstream sequences in vitro. Previous in vivo studies suggested that efficient polyadenylation at the HIV-1 poly(A) signal requires a spacing of at least 250 nucleotides between the 5' cap site and the AAUAAA. Our in vitro analyses indicated that a precursor containing the defined upstream and downstream sequences was efficiently cleaved at the polyadenylation site when the distance between the 5' cap and the AAUAAA was reduced to at least 140 nucleotides, which is less than the distance predicted from in vivo studies. This cleavage was dependent on the presence of the upstream element.

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The efficiency of RNA 3'-end formation is determined by the distance between the cap site and the poly(A) site in spleen necrosis virus.

Cited by 45 publications

References 30 publications

Relative roles of signals upstream of AAUAAA and promoter proximity in regulation of human immunodeficiency virus type 1 mRNA 3' end formation.

Relative roles of signals upstream of AAUAAA and promoter proximity in regulation of human immunodeficiency virus type 1 mRNA 3' end formation.

Effects of Homology Length in the Repeat Region on Minus-Strand DNA Transfer and Retroviral Replication

Elements upstream of the AAUAAA within the human immunodeficiency virus polyadenylation signal are required for efficient polyadenylation in vitro.

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