Search PubMed⌕ Search

Biomedical subjects

G Khoury

Publications and source records attributed to G Khoury.

At least 127 records · Page 7Linked to original sources
PubMed1980

A rapid enzymatic DNA sequencing technique: determination of sequence alterations in early simian virus 40 temperature sensitive and deletion mutants.

We have adapted a rapid sequencing technique from the enzymatic nick-translation method of Maat and Smith. The Forward-Backward procedure employs both synthetic and 3' to 5' exonucleolytic activities of E. coli DNA polymerase I to achieve greater reliability, especially in reading stretches of the same nucleotide. The technique has been employed to determine sequence alterations in four early SV40 temperature-sensitive (tsA) point mutants and five early SV40 viable deletion mutants. The nucleotide sequence of these mutants provides an insight into their biological properties.

Amino Acid Sequence↗
PubMed1980

A small RNA induced late in simian virus 40 infection can associate with early viral mRNAs.

Analysis of total cytoplasmic and polyadenylylated cytoplasmic RNA from cells lytically infected with simian virus 40 (SV40) has demonstrated the presence of a small RNA, approximately 65 nucleotides long, that is induced late in lytic infection. This small RNA is apparently specific in size and sequence and is not selected on columns of oligo(dT)-cellulose. It is homologous to a region of the early SV40 mRNAs (and to the late DNA strand), starting approximately 250 nucleotides from the 3' end of the early mRNAs (SV40 map position 0.21). The function of this RNA in the viral cycle and its source are unknown at this time; however, its temporal expression, unique sequence, and interesting region of homology within the SV40 genome suggest a possible role in the control of SV40 gene expression.

Animals↗
PubMed1980

Effect of a tsA mutation of simian virus 40 late gene expression: variations between host cell lines.

Infection of AGMK or CV-1 cells by the early simian virus 40 mutant tsA58 at the permissive temperature (32 degrees C) followed by a shift to the nonpermissive temperature (41 degrees C) caused a substantial decrease in the levels of late viral RNA in the cytoplasm of AGMK cells but not CV-1 cells. At the translational level, this depression of late viral RNA levels was reflected by a decrease in late viral protein synthesis. Thus, in AGMK cells, an early region gene product (presumably large T-antigen) appeared to be continuously required for efficient expression of the late viral genes. In contrast, late simian virus 40 gene expression, once it is initiated in CV-1 cells, continued efficiently regardless of the tsA mutation. The difference in expression of the late simian virus 40 genes in these tsA mutant-infected monkey kidney cell lines may reflect a difference in host cell proteins which regulate viral gene expression in conjunction with early viral proteins.

Animals↗
PubMed1980

Adenovirus regulation of simian virus 40 macromolecular synthesis.

At low multiplicities of infection (1 to 5 PFU/cell), human adenoviruses grew inefficiently in monkey kidney cells as the result of a late transcriptional and/or translational defect. Through an unknown mechanism, coinfection with simian virus 40 (SV40) could overcome this block. Coincident with the onset of adenovirus DNA replication in coinfected monkey cells, however, there was a marked inhibition of SV40 DNA replication. Since this inhibition in SV40 DNA synthesis occurred in the presence of normal SV40 transcription and protein synthesis, we suggest that adenovirus competes with SV40 for molecules specifically required for DNA synthesis (e.g., SV40 T-antigen). At high multiplicities of adenovirus infection (50 PFU/cell), adenoviruses could grow efficiently in monkey kidney cells. In cells coinfected under these conditions, SV40 transcription was generally inhibited, as shown by a decrease in nuclear and cytoplasmic SV40 mRNA.

Adenoviruses, Human↗
PubMed1980

Control of simian virus 40 gene expression at the levels of RNA synthesis and processing: thermally induced changes in the ratio of the simian virus 40 early mRNA's and proteins.

Examination of the simian virus 40 early mRNA's from infected AGMK or CV-1 cells showed that the ratio of large T- to small t-antigen mRNA's increased with an increased incubation temperature. In tsA58 mutant-infected cells, an increased incubation temperature resulted in the overproduction of early RNAs'; however, the ratio of the early mRNA's was the same, at any temperature, in both wild-type- and tsA58-infected cells. Thus, the thermally induced alteration in the early mRNA ratios was apparently not affected by the tsA mutation or by the overproduction of early RNA in tsA mutant-infected cells. Time course studies at various temperatures showed that, although the ratio of large T- to small t-antigen mRNA's increased with temperature, at any one temperature it was consistent from early to late times of infection. Furthermore, the ratio of the early mRNA's adjusted in temperature shift experiments. Thus, the ratio of the early mRNA's appeared to be intrinsic to the thermodynamic environment of the cell. The thermally induced alterations in the early mRNA's were reflected at the protein level by parallel changes in the ratio of large T- to small t-antigens. These data suggest a level of gene expression control which may function at the stage of splicing.

Animals↗
PubMed1980

New chimeric splice junction in adenovirus type 2-simian virus 40 hybrid viral mRNA.

We have examined hybrid viral RNAs synthesized in both human and monkey cells infected by three nondefective adenovirus type 2 (Ad2)-simian virus 40 (SV40) hybrid viruses; Ad2+ND1, Ad2+ND2, and Ad2+ND4. Most of the hybrid viral RNA molecules appeared to be initiated within adenoviral sequences, but were polyadenylated on their 3' end at the early SV40 mRNA polyadenylation site. The Ad2+ND4 stock of virus was not homogeneous, but consisted of two principle populations of viral DNA. Both populations contained a segment of SV40 DNA extending from the SV40 map positions 0.63 to 0.11 in a left-to-right orientation at adenovirus map position 0.82. One population contained an intact SV40 segment, whereas the other (representing 80 to 85% of the population) has a 500-base pair deletion mapping from approximately 0.60 to 0.50 SV40 map units. This deletion encompassed the SV40 DNA segment which encodes the early SV40 splice sites. Cells infected by the mixed Ad2+ND4 population induced the synthesis of at least three major SV40 RNA species among the hybrid viral transcripts. The most abundant of these hybrid mRNA's appeared only late in the lytic cycle, after the onset of viral DNA replication. It contained an RNA splice junction which extended from a donor (5') nucleotide within the adenoviral RNA sequences to an acceptor (3') splice site within the early region of SV40 at 0.46 SV40 map units. This SV40 acceptor splice site was remarkable in that its use has not been detected in the spliced viral mRNA's of SV40-infected or -transformed cells.

Adenoviridae↗
PubMed1980

Simian virus 40-associated small RNA: mapping on the simian virus 40 genome and characterization of its synthesis.

The simian virus 40 (SV40)-associated small RNA (SAS-RNA) has previously been shown to arise late in SV40 lytic infection and to bear homology with the SV40 early mRNA's, or the SV40 genome, at map position 0.21. By using hybridization analysis, we determined that the SAS-RNA is between 62 and 65 nucleotides in length and its homology region lies between nucleotides 2760 and 2825 of the SV40 late(+) DNA strand. Viable deletion mutants which lacked part or all of this region made no SAS-RNA, strongly indicating that this is the coding region of the SAS-RNA. The expected sequence for the SAS-RNA, determined from the DNA sequence between nucleotides 2760 and 2825, appeared to be very pyrimidine rich (76% uridine and cytidine). Deletion or alteration of sequences immediately preceding the SAS-RNA coding region (approximately nucleotides 2716 to 2748) resulted in the loss of SAS-RNA production. These sequences may be part of a promotor for SAS-RNA synthesis or a processing site for its excision from long nuclear late transcripts. Under growth conditions where late transcription was not fully initiated (tsA58 at 41 degrees C; wild-type SV40 in the presence of 1-beta-D-arabinofuranosylcytosine), no SAS-RNA was produced, indicating that the expression of the SAS-RNA is regulated by a mechanism related to the control of late transcription.

Base Sequence↗
PubMed1979

BKV splice sequences based on analysis of preferred donor and acceptor sites.

We have determined the DNA sequences which correspond to the splicing regions in the transcripts of human papovavirus BKV, an evolutionary variant of SV40. To precisely localize the excision points in the BKV sequence, we have conducted a preliminary analysis of numerous viral and eukaryotic splice site sequences. This analysis suggests that the preferred sequence for the donor site belongs to at least one of four groups: Pu↓GTAxG, Pu↓GTAxxT, Pu↓GTxxGT, Pu↓GTxAG (↓ = the cleavage site). These four groups derive from the two basic sequences, Pu↓GTxxG and Pu↓GTA. An optimal donor site might be: AG↓GTAAGT. The preferred sequence for the acceptor site is of the form PyPyxPyAG↓; no dinucleotide AG occurs within 13 nucleotides prior to the terminal AG of the 3' end of the intervening sequences. As we have found in this study of BKV splice sites, the sequences for the preferred donor and acceptor sites provide predictive value in localizing RNA splice points when the DNA sequence is known.

BK Virus↗
PubMed1979

The genome of human papovavirus BKV.

The complete DNA sequence of human papovavirus BKV(Dun), consisting of 5153 nucleotide pairs, is presented. We describe the segments of the genome which correspond to the replication origin, the tandem repeated sequences, the 5' and 3' ends of the mRNAs, the splice sites, the early and late viral proteins and the putative viral polypeptides. These BKV DNA sequences are compared with analogous regions in the SV40 and Py virus genomes in an attempt to localize viral functions for lytic growth and transformation.

Amino Acid Sequence↗
PubMed1979

Deletion mutants of simian virus 40 defective in biosynthesis of late viral mRNA.

Deletion mutants of simian virus 40 defective in late gene functions have been examined for genetic elements that control the biosynthesis of the viral mRNAs. Mutant-specific RNA from infected cells was identified by CH3Hg/(OH)/agarose gel electrophoresis and mapped by the nuclease S1 technique. Altered RNA species, shortened by a size equivalent to the deletion region, can be detected in cells infected with these mutants lacking sequences within the body RNA segments. On the other hand, mutants that lack the 5'-end of the body sequences (a splice junction) fail to accumulate the respective shortened RNA species. In particular, mutant dl-2301 whose deletion includes both the leader and body splice junctions plus the intervening sequences, exhibits a polar effect on a distal gene. Whereas the late region of dl-2301 can be transcribed normally, the mutant defect appears to be associated with little or no accumulation of the mutant-specific late RNA in the infected cells. These results suggest that splice junctions or the intervening sequences, or both, in the viral genome are control signals for post-transcriptional processing of the viral RNA.

Chromosome Deletion↗