The fission yeast Schizosaccharomyces pombe has two distinct tRNase ZLs encoded by two different genes and differentially targeted to the nucleus and mitochondria

2011 ◽  
Vol 435 (1) ◽  
pp. 103-111 ◽  
Author(s):  
Xuhua Gan ◽  
Jing Yang ◽  
Jun Li ◽  
Haiyan Yu ◽  
Hongmei Dai ◽  
...  
Keyword(s):  
Schizosaccharomyces Pombe ◽  
Fission Yeast ◽  
Mitochondrial Protein ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
Yeast Saccharomyces Cerevisiae ◽  
Lysine Residues ◽  
Trnase Z ◽  
High Level

tRNase Z is the endonuclease that is involved in tRNA 3′-end maturation by removal of the 3′-trailer sequences from tRNA precursors. Most eukaryotes examined to date, including the budding yeast Saccharomyces cerevisiae and humans, have a single long form of tRNase Z (tRNase ZL). In contrast, the fission yeast Schizosaccharomyces pombe contains two candidate tRNase ZLs encoded by the essential genes sptrz1+ and sptrz2+. In the present study, we have expressed recombinant SpTrz1p and SpTrz2p in S. pombe. Both recombinant proteins possess precursor tRNA 3′-endonucleolytic activity in vitro. SpTrz1p localizes to the nucleus and has a simian virus 40 NLS (nuclear localization signal)-like NLS at its N-terminus, which contains four consecutive arginine and lysine residues between residues 208 and 211 that are critical for the NLS function. In contrast, SpTrz2p is a mitochondrial protein with an N-terminal MTS (mitochondrial-targeting signal). High-level overexpression of sptrz1+ has no detectable phenotypes. In contrast, strong overexpression of sptrz2+ is lethal in wild-type cells and results in morphological abnormalities, including swollen and round cells, demonstrating that the correct expression level of sptrz2+ is critical. The present study provides evidence for partitioning of tRNase Z function between two different proteins in S. pombe, although we cannot rule out specialized functions for each protein.

scholarly journals Sequence-specific interaction of histones with the simian virus 40 enhancer region in vitro.

1985 ◽  
Vol 260 (23) ◽  
pp. 12394-12397
Author(s):  
M F Clarke ◽  
P C FitzGerald ◽  
J M Brubaker ◽  
R T Simpson
Keyword(s):  
Specific Interaction ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
Enhancer Region

scholarly journals Stimulation of the Human Heat Shock Protein 70 Promoter in Vitro by Simian Virus 40 Large T Antigen

1989 ◽  
Vol 264 (27) ◽  
pp. 16160-16164
Author(s):  
I C Taylor ◽  
W Solomon ◽  
B M Weiner ◽  
E Paucha ◽  
M Bradley ◽  
...  
Keyword(s):  
Heat Shock ◽  
Heat Shock Protein ◽  
Heat Shock Protein 70 ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
T Antigen ◽  
Large T Antigen ◽  
Human Heat Shock Protein ◽  
Stimulation Of

Intrinsic mineralization defect inHyp mouse osteoblasts

1998 ◽  
Vol 275 (4) ◽  
pp. E700-E708 ◽  
Author(s):  
Z. S. Xiao ◽  
M. Crenshaw ◽  
R. Guo ◽  
T. Nesbitt ◽  
M. K. Drezner ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Human Disease ◽  
Normal Mouse ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
Diffusible Factor ◽  
Murine Homologue ◽  
Mineralization Defect ◽  
Inactivating Mutations

X-linked hypophosphatemia (XLH) is caused by inactivating mutations of PEX, an endopeptidase of uncertain function. This defect is shared by Hyp mice, the murine homologue of the human disease, in which a 3′ Pex deletion has been documented. In the present study, we report that immortalized osteoblasts derived from the simian virus 40 (SV40) transgenic Hyp mouse (TMOb- Hyp) have an impaired capacity to mineralize extracellular matrix in vitro. Compared with immortalized osteoblasts from the SV40 transgenic normal mouse (TMOb-Nl), osteoblast cultures from the SV40 Hyp mouse exhibit diminished 45Ca accumulation into extracellular matrix (37 ± 6 vs. 1,484 ± 68 counts ⋅ min−1 ⋅ μg protein−1) and reduced formation of mineralization nodules. Moreover, in coculture experiments, we found evidence that osteoblasts from the SV40 Hyp mouse produce a diffusible factor that blocks mineralization of extracellular matrix in normal osteoblasts. Our findings indicate that abnormal PEX in osteoblasts is associated with the accumulation of a factor(s) that inhibits mineralization of extracellular matrix in vitro.

scholarly journals Analysis of a transgenic mouse containing simian virus 40 and v-myc sequences.

1985 ◽  
Vol 5 (4) ◽  
pp. 642-648 ◽  
Author(s):  
J A Small ◽  
D G Blair ◽  
S D Showalter ◽  
G A Scangos
Keyword(s):  
Cell Lines ◽  
Choroid Plexus Papilloma ◽  
Simian Virus 40 ◽  
Soft Agar ◽  
Simian Virus ◽  
T Antigen ◽  
Primary Cell ◽  
Tissue Samples ◽  
Primary Cell Lines

Two plasmids, one containing the simian virus 40 (SV40) genome and the mouse metallothionein I gene and one containing the v-myc gene of avian myelocytomatosis virus MC29, were coinjected into mouse embryos. Of the 13 surviving mice, one, designated M13, contained both myc and SV40 sequences. This mouse developed a cranial bulge identified as a choroid plexus papilloma at 13 weeks and was subsequently sacrificed; tissue samples were taken for further analysis. Primary cell lines derived from these tissues contained both myc and SV40 DNA. No v-myc mRNA could be detected, although SV40 mRNA was present in all of the cell lines tested. T antigen also was expressed in all of the cell lines analyzed. These data suggest that SV40 expression was involved in the abnormalities of mouse M13 and was responsible for the transformed phenotype of the primary cell lines. Primary cell lines from this mouse were atypical in that the population rapidly became progressively more transformed with time in culture based on the following criteria: morphology, growth rate, and the ability to grow in soft agar and in serum-free medium. The data also suggest that factors present in the mouse regulated the ability of SV40 to oncogenically transform most cells and that in vitro culture of cells allowed them to escape those factors.

The AAUAAA sequence is required both for cleavage and for polyadenylation of simian virus 40 pre-mRNA in vitro

1986 ◽  
Vol 6 (7) ◽  
pp. 2317-2323
Author(s):  
D Zarkower ◽  
P Stephenson ◽  
M Sheets ◽  
M Wickens
Keyword(s):  
Hela Cell ◽  
Simian Virus 40 ◽  
Nuclear Extract ◽  
Simian Virus ◽  
Polyadenylation Site ◽  
Single Base ◽  
Cleavage And Polyadenylation ◽  
Cell Nuclear Extract

The sequence AAUAAA is found near the polyadenylation site of eucaryotic mRNAs. This sequence is required for accurate and efficient cleavage and polyadenylation of pre-mRNAs in vivo. In this study we show that synthetic simian virus 40 late pre-mRNAs are cleaved and polyadenylated in vitro in a HeLa cell nuclear extract, and that cleavage in vitro is abolished by each of four different single-base changes in AAUAAA. In this same extract, precleaved RNAs (RNAs with 3' termini at the polyadenylation site) are efficiently polyadenylated. This in vitro polyadenylation reaction also requires the AAUAAA sequence.

Replication of cloned DNA containing the Alu family sequence during cell extract-promoting simian virus 40 DNA synthesis

1984 ◽  
Vol 4 (8) ◽  
pp. 1476-1482
Author(s):  
H Ariga
Keyword(s):  
Dna Replication ◽  
Dna Synthesis ◽  
Rna Polymerase Iii ◽  
Simian Virus 40 ◽  
Cell Extract ◽  
Simian Virus ◽  
T Antigen ◽  
Sv40 T Antigen ◽  
Sv40 Dna

The replicating activity of several cloned DNAs containing putative origin sequences was examined in a cell-free extract that absolutely depends on simian virus 40 (SV40) T antigen promoting initiation of SV40 DNA replication in vitro. Of the three DNAs containing the human Alu family sequence (BLUR8), the origin of (Saccharomyces cerevisiae plasmid 2 micron DNA (pJD29), and the yeast autonomous replicating sequence (YRp7), only BLUR8 was active as a template. Replication in a reaction mixture with BLUR8 as a template was semiconservative and not primed by a putative RNA polymerase III transcript synthesized on the Alu family sequence in vitro. Pulse-chase experiments showed that the small-sized DNA produced in a short-term incubation was converted to full-length closed circular and open circular DNAs in alkaline sucrose gradients. DNA synthesis in extracts began in a region of the Alu family sequence and was inhibited 80% by the addition of anti-T serum. Furthermore, partially purified T antigen bound the Alu family sequence in BLUR8 by the DNA-binding immunoassay. These results suggest that SV40 T antigen recognizes the Alu family sequence, similar to the origin sequence of SV40 DNA, and initiates semiconservative DNA replication in vitro.

Carcinogen-induced DNA amplification in vitro: overreplication of the simian virus 40 origin region in extracts from carcinogen-treated CO60 cells

1990 ◽  
Vol 10 (1) ◽  
pp. 75-83
Author(s):  
Y Berko-Flint ◽  
S Karby ◽  
D Hassin ◽  
S Lavi
Keyword(s):  
De Novo ◽  
Chinese Hamster ◽  
Simian Virus 40 ◽  
Simian Virus ◽  
T Antigen ◽  
Dna Molecules ◽  
Viral Origin ◽  
Cell Extracts ◽  
Origin Region

An in vitro system to study carcinogen-induced amplification in simian virus 40 (SV40)-transformed Chinese hamster (CO60) cells is described. SV40 amplification in this system resembled in many aspects the viral overreplication observed in drug-treated CO60 cells. Cytosolic extracts from N-methyl-N'-nitro-N-nitrosoguanidine-treated cells supported de novo DNA synthesis in the presence of excess exogenous T antigen and the SV40-containing plasmid pSVK1. The pattern of viral replication in these extracts was unique, since only the 2.4-kilobase-pair region spanning the origin was overreplicated, whereas distal sequences were not replicated significantly. Extracts from control cells supported only marginal levels of replication. In HeLa extracts, complete SV40 DNA molecules were replicated efficiently. The overreplication of the origin region in CO60 cell extracts was bidirectional and symmetrical. A fraction of the newly synthesized DNA molecules underwent a second round of replication, yielding MboI-sensitive fragments representing the 2.4-kilobase-pair region around the origin. The mechanisms controlling the amplification of the viral origin region, the nature of the cellular factors induced in the carcinogen-treated cells, and their putative association with general drug-induced SOS-like responses are discussed.

Site-specific mutagenesis of cdc2+, a cell cycle control gene of the fission yeast Schizosaccharomyces pombe

1986 ◽  
Vol 6 (10) ◽  
pp. 3523-3530
Author(s):  
R Booher ◽  
D Beach
Keyword(s):  
Schizosaccharomyces Pombe ◽  
Fission Yeast ◽  
Cell Cycle Control ◽  
Temperature Sensitive ◽  
Protein Kinase Activity ◽  
Atp Binding Site ◽  
Leu2 Gene ◽  
Cdc2 Gene ◽  
A Site

The cdc2+ gene of Schizosaccharomyces pombe is homologous to the CDC28 gene of Saccharomyces cerevisiae. Both genes share limited homology with vertebrate protein kinases and have protein kinase activity. cdc2+ has been subjected to mutagenesis in vitro. A null allele of the gene, constructed by insertion of the S. cerevisiae LEU2 gene into a site within the gene, has a phenotype similar to that of many temperature-sensitive alleles of cdc2. Mutations within the predicted ATP-binding site and in a region which may be a site of phosphorylation result in loss of cdc2+ activity. A single substitution of Gly-146 to Asp-146 has been identified in cdc2-1w, a dominant activated allele of the gene. The four introns within the cdc2+ gene have been deleted. The resulting gene not only functions in fission yeast but also rescues cdc28(Ts) strains of S. cerevisiae, a property which is not shared by the genomic cdc2+ gene.

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