scholarly journals The number of ribosomes on simian virus 40 late 16S mRNA is determined in part by the nucleotide sequence of its leader.

1984 ◽  
Vol 4 (4) ◽  
pp. 813-816 ◽  
Author(s):  
A Barkan ◽  
J E Mertz
Keyword(s):  
Nucleotide Sequence ◽  
Direct Evidence ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
The Body ◽  
Size Distributions ◽  
Wild Type ◽  
Leader Protein ◽  
16S Rna

The size distributions of polyribosomes containing each of three simian virus 40 late 16S mRNA species that differ in nucleotide sequence only within their leaders were determined. The two 16S RNA species with shorter leaders were incorporated into polysomes that were both larger (on average) and narrower in size distribution than was the predominant wild-type 16S RNA. Therefore, the nucleotide sequence of the leader can influence the number of ribosomes present on the body of an mRNA molecule. We propose a model in which the excision from leaders of sizeable translatable regions permits more frequent utilization of internally located translation initiation signals, thereby enabling genes encoded within the bodies of polygenic mRNAs to be translated at higher rates. In addition, the data provide the first direct evidence that VP1 can, indeed, be synthesized in vivo from the species of 16S mRNA that also encodes the 61-amino acid leader protein.

The number of ribosomes on simian virus 40 late 16S mRNA is determined in part by the nucleotide sequence of its leader

1984 ◽  
Vol 4 (4) ◽  
pp. 813-816
Author(s):  
A Barkan ◽  
J E Mertz
Keyword(s):  
Nucleotide Sequence ◽  
Direct Evidence ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
The Body ◽  
Size Distributions ◽  
Wild Type ◽  
Leader Protein ◽  
16S Rna

The size distributions of polyribosomes containing each of three simian virus 40 late 16S mRNA species that differ in nucleotide sequence only within their leaders were determined. The two 16S RNA species with shorter leaders were incorporated into polysomes that were both larger (on average) and narrower in size distribution than was the predominant wild-type 16S RNA. Therefore, the nucleotide sequence of the leader can influence the number of ribosomes present on the body of an mRNA molecule. We propose a model in which the excision from leaders of sizeable translatable regions permits more frequent utilization of internally located translation initiation signals, thereby enabling genes encoded within the bodies of polygenic mRNAs to be translated at higher rates. In addition, the data provide the first direct evidence that VP1 can, indeed, be synthesized in vivo from the species of 16S mRNA that also encodes the 61-amino acid leader protein.

scholarly journals A splice junction deletion deficient in the transport of RNA does not polyadenylate nuclear RNA.

1983 ◽  
Vol 3 (8) ◽  
pp. 1381-1388 ◽  
Author(s):  
L P Villarreal ◽  
R T White
Keyword(s):  
Simian Virus 40 ◽  
Splice Junction ◽  
Simian Virus ◽  
The Body ◽  
Splice Acceptor Site ◽  
Acceptor Site ◽  
Wild Type ◽  
Normal Position ◽  
Nuclear Rna ◽  
Late Region

A late region deletion mutant of simian virus 40 (dl5) was previously shown to be deficient in the transport of nuclear RNA. This is a splice junction deletion that has lost the 3' end of an RNA leader, an intervening sequence, and the 5' end of the splice acceptor site on the body of the mRNA. In this report, we analyzed the steady-state structure of the untransported nuclear RNA. The 5' ends of this RNA are heterogeneous but contain a prominent 5' end at the normal position (nucleotide 325) in addition to several other prominent 5' ends not seen in wild-type RNA. The 3' end of this RNA does not occur at the usual position (nucleotide 2674) of polyadenylation; instead, this RNA is non-polyadenylated, with the 3' end occurring either downstream or upstream of the normal position.

scholarly journals Specific protein binding to the simian virus 40 enhancer in vitro.

1986 ◽  
Vol 6 (6) ◽  
pp. 2098-2105 ◽  
Author(s):  
A G Wildeman ◽  
M Zenke ◽  
C Schatz ◽  
M Wintzerith ◽  
T Grundström ◽  
...  
Keyword(s):  
Point Mutations ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
Specific Protein ◽  
Dnase I ◽  
Wild Type ◽  
Dnase I Footprinting ◽  
Nuclear Extracts

HeLa cell nuclear extracts and wild-type or mutated simian virus 40 enhancer DNA were used in DNase I footprinting experiments to study the interaction of putative trans-acting factors with the multiple enhancer motifs. We show that these nuclear extracts contain proteins that bind to these motifs. Because point mutations which are detrimental to the activity of a particular enhancer motif in vivo specifically prevent protection of that motif against DNase I digestion in vivo, we suggest that the bound proteins correspond to trans-acting factors involved in enhancement of transcription. Using mutants in which the two domains A and B of the simian virus 40 enhancer are either separated by insertion of DNA fragments or inverted with respect to their natural orientation, we also demonstrate that the trans-acting factors bind independently to the two domains.

RNA sequence containing hexanucleotide AAUAAA directs efficient mRNA polyadenylation in vitro

1985 ◽  
Vol 5 (2) ◽  
pp. 373-379
Author(s):  
J L Manley ◽  
H Yu ◽  
L Ryner
Keyword(s):  
Nucleotide Sequence ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
Whole Cell Lysate ◽  
Cell Lysate ◽  
Polyadenylic Acid ◽  
Specific Nucleotide Sequence ◽  
Specific Nucleotide

To determine whether a specific nucleotide sequence is required to direct polyadenylation of a simian virus 40 early pre-mRNA in a soluble HeLa whole-cell lysate, we constructed a series of rearranged and deleted DNA templates, transcribed them in vitro, and determined whether the resultant RNAs could be polyadenylated when incubated in whole-cell lysate. When a 237-base-pair DNA fragment encoding the 3' end of the simian virus 40 early pre-mRNA was transferred to recombinant plasmids encoding RNAs that were not substrates for polyadenylation, the resultant RNAs could now be polyadenylated efficiently. In one case, the chimeric RNA was polyadenylated even more efficiently than was the original simian virus 40 early transcript. Analysis of the RNAs produced from the deletion mutant templates revealed that only RNAs containing at least one copy of the AAUAAA sequence situated near the 3' end and implicated in 3'-end formation and polyadenylation in vivo could be polyadenylated in vitro. Surprisingly, this sequence directed polyadenylation of pre-mRNAs not only when near the RNA 3' end, i.e., 50 nucleotides or less away, but also when the 3' end was situated over 400 nucleotides downstream. Thus, our results show that a polyadenylic acid polymerase activity in HeLa lysates can recognize a specific nucleotide sequence in pre-mRNA and then, in the absence of the nucleolytic cleavage that presumably occurs in vivo, locate the RNA 3' end and use it as a primer for polyadenylic acid synthesis.

scholarly journals RNA sequence containing hexanucleotide AAUAAA directs efficient mRNA polyadenylation in vitro.

1985 ◽  
Vol 5 (2) ◽  
pp. 373-379 ◽  
Author(s):  
J L Manley ◽  
H Yu ◽  
L Ryner
Keyword(s):  
Nucleotide Sequence ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
Whole Cell Lysate ◽  
Cell Lysate ◽  
Polyadenylic Acid ◽  
Specific Nucleotide Sequence ◽  
Specific Nucleotide

To determine whether a specific nucleotide sequence is required to direct polyadenylation of a simian virus 40 early pre-mRNA in a soluble HeLa whole-cell lysate, we constructed a series of rearranged and deleted DNA templates, transcribed them in vitro, and determined whether the resultant RNAs could be polyadenylated when incubated in whole-cell lysate. When a 237-base-pair DNA fragment encoding the 3' end of the simian virus 40 early pre-mRNA was transferred to recombinant plasmids encoding RNAs that were not substrates for polyadenylation, the resultant RNAs could now be polyadenylated efficiently. In one case, the chimeric RNA was polyadenylated even more efficiently than was the original simian virus 40 early transcript. Analysis of the RNAs produced from the deletion mutant templates revealed that only RNAs containing at least one copy of the AAUAAA sequence situated near the 3' end and implicated in 3'-end formation and polyadenylation in vivo could be polyadenylated in vitro. Surprisingly, this sequence directed polyadenylation of pre-mRNAs not only when near the RNA 3' end, i.e., 50 nucleotides or less away, but also when the 3' end was situated over 400 nucleotides downstream. Thus, our results show that a polyadenylic acid polymerase activity in HeLa lysates can recognize a specific nucleotide sequence in pre-mRNA and then, in the absence of the nucleolytic cleavage that presumably occurs in vivo, locate the RNA 3' end and use it as a primer for polyadenylic acid synthesis.

scholarly journals Mutual Balance of Histone Deacetylases 1 and 2 and the Acetyl Reader ATAD2 Regulates the Level of Acetylation of Histone H4 on Nascent Chromatin of Human Cells

2020 ◽  
Vol 40 (9) ◽  
Author(s):  
Pavlo Lazarchuk ◽  
John Hernandez-Villanueva ◽  
Maria N. Pavlova ◽  
Alexander Federation ◽  
Michael MacCoss ◽  
...  
Keyword(s):  
Histone Deacetylases ◽  
De Novo ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
Histone H4 ◽  
Wild Type ◽  
Histone Deacetylase 1 ◽  
Replication Fork Progression ◽  
Model Cell

ABSTRACT Newly synthesized histone H4 that is incorporated into chromatin during DNA replication is acetylated on lysines 5 and 12. Histone deacetylase 1 (HDAC1) and HDAC2 are responsible for reducing H4 acetylation as chromatin matures. Using CRISPR-Cas9-generated hdac1- or hdac2-null fibroblasts, we determined that HDAC1 and HDAC2 do not fully compensate for each other in removing de novo acetyls on H4 in vivo. Proteomics of nascent chromatin and proximity ligation assays with newly replicated DNA revealed the binding of ATAD2, a bromodomain-containing posttranslational modification (PTM) reader that recognizes acetylated H4. ATAD2 is a transcription facilitator overexpressed in several cancers and in the simian virus 40 (SV40)-transformed human fibroblast model cell line used in this study. The recruitment of ATAD2 to nascent chromatin was increased in hdac2 cells over the wild type, and ATAD2 depletion reduced the levels of nascent chromatin-associated, acetylated H4 in wild-type and hdac2 cells. We propose that overexpressed ATAD2 shifts the balance of H4 acetylation by protecting this mark from removal and that HDAC2 but not HDAC1 can effectively compete with ATAD2 for the target acetyls. ATAD2 depletion also reduced global RNA synthesis and nascent DNA-associated RNA. A moderate dependence on ATAD2 for replication fork progression was noted only for hdac2 cells overexpressing the protein.

Specific protein binding to the simian virus 40 enhancer in vitro

1986 ◽  
Vol 6 (6) ◽  
pp. 2098-2105
Author(s):  
A G Wildeman ◽  
M Zenke ◽  
C Schatz ◽  
M Wintzerith ◽  
T Grundström ◽  
...  
Keyword(s):  
Point Mutations ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
Specific Protein ◽  
Dnase I ◽  
Wild Type ◽  
Dnase I Footprinting ◽  
Nuclear Extracts

HeLa cell nuclear extracts and wild-type or mutated simian virus 40 enhancer DNA were used in DNase I footprinting experiments to study the interaction of putative trans-acting factors with the multiple enhancer motifs. We show that these nuclear extracts contain proteins that bind to these motifs. Because point mutations which are detrimental to the activity of a particular enhancer motif in vivo specifically prevent protection of that motif against DNase I digestion in vivo, we suggest that the bound proteins correspond to trans-acting factors involved in enhancement of transcription. Using mutants in which the two domains A and B of the simian virus 40 enhancer are either separated by insertion of DNA fragments or inverted with respect to their natural orientation, we also demonstrate that the trans-acting factors bind independently to the two domains.

A splice junction deletion deficient in the transport of RNA does not polyadenylate nuclear RNA

1983 ◽  
Vol 3 (8) ◽  
pp. 1381-1388
Author(s):  
L P Villarreal ◽  
R T White
Keyword(s):  
Simian Virus 40 ◽  
Splice Junction ◽  
Simian Virus ◽  
The Body ◽  
Splice Acceptor Site ◽  
Acceptor Site ◽  
Wild Type ◽  
Normal Position ◽  
Nuclear Rna ◽  
Late Region

A late region deletion mutant of simian virus 40 (dl5) was previously shown to be deficient in the transport of nuclear RNA. This is a splice junction deletion that has lost the 3' end of an RNA leader, an intervening sequence, and the 5' end of the splice acceptor site on the body of the mRNA. In this report, we analyzed the steady-state structure of the untransported nuclear RNA. The 5' ends of this RNA are heterogeneous but contain a prominent 5' end at the normal position (nucleotide 325) in addition to several other prominent 5' ends not seen in wild-type RNA. The 3' end of this RNA does not occur at the usual position (nucleotide 2674) of polyadenylation; instead, this RNA is non-polyadenylated, with the 3' end occurring either downstream or upstream of the normal position.

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