PubMed Health⌕ Search

Biomedical subjects

R Levis

Publications and source records attributed to R Levis.

At least 37 records · Page 2Linked to original sources
PubMed1984

Transformation of white locus DNA in drosophila: dosage compensation, zeste interaction, and position effects.

P-element-mediated DNA transformation was used to generate transformants carrying segments of DNA from the white locus of D. melanogaster. The vast majority of transduced copies of an 11.7 or a 14.3 kb segment of DNA from white successfully rescued the white- eye-color phenotype when inserted in many different chromosomal locations. However, two transformants with abnormal eye pigmentation--apparently a consequence of the genomic positions of the transduced white gene--were also recovered. In all seven cases tested, autosomal insertions of white, which is dosage-compensated in its normal location on the X chromosome, retained the property of dosage compensation. In contrast to the relative insensitivity of eye-color pigmentation and dosage compensation to genomic position, the transduced white DNA segments differed widely in their interactions with the zeste mutation, ranging from greater than normal repression by zeste to insensitivity to the presence of zeste.

Animals↗
PubMed1984

Effects of transposable element insertions on RNA encoded by the white gene of Drosophila.

We have examined the manner in which transposable element insertions affect the expression of the white gene of Drosophila by analyzing polyadenylated RNA of flies with each of nine insertions in or near the gene. In five mutants having insertions in the transcribed sequences of white, transcripts initiating at the white promoter are truncated within the insertions. Two insertions in the 3 kb intron of white alter neither the amount nor the structure of the mature white RNA. An insertion near the 5' end of the gene blocks the accumulation of any white transcripts. Another insertion, located 1.2 kb upstream from the transcribed region of the gene, causes a mutant phenotype yet surprisingly has no obvious effect on the structure or abundance of the major white RNA. We also show that a mutation at each of two other loci that modulate the phenotype of the white-apricot insertion mutant are correlated with small but significant changes in the pattern of white transcripts.

Animals↗
PubMed1983

Transcription of the white locus in Drosophila melanogaster.

Genetic studies of the white locus have shown that it has a distal region where structural mutations occur and a proximal region where regulatory mutations occur. To better understand the molecular basis of this genetic organization we have analyzed white locus transcription. A 2.7-kilobase transcript comprising 0.0005% of poly(A)-RNA was detected in RNA prepared from pupae or adults. The structure of this transcript helps clarify some unusual genetic properties of the locus. There is a small 5' exon separated from the majority of the sequences found in the mature RNA by an intron of approximately 2.8 kilobases. This 5' exon is from the proximal region of the locus, whereas the main body of the RNA maps to the distal region. The mutationally silent region between the proximal and distal regions corresponds to the large intron. We have identified the family and determined the exact location of a number of transposable element insertions within the locus. These results show that transposable element insertions within introns can be without phenotypic effect. We have also investigated the effect on the white transcript of the zeste mutation, which represses white locus expression as judged by eye color phenotype. The RNA was unchanged in size or abundance in poly(A)-RNA from adult flies. This demonstrates that the zeste-white interaction does not occur by simply repressing transcription of the white locus in all tissues.

Journal Article↗
PubMed1983

Evolution of virus and defective-interfering RNAs in BHK cells persistently infected with Sindbis virus.

We analyzed a BHK cell line persistently infected with Sindbis virus for 16 months and a virus (Sin-16) cloned from these cells. Sin-16 virus was resistant to the defective interfering particles present in the original infection. We found that (i) cells infected with Sin-16 were impaired in the processing of a viral precursor glycoprotein, (ii) high-multiplicity passaging of Sin-16 gave rise to a variant that was able to generate and be inhibited by defective-interfering particles to which the original Sin-16 virus was resistant, and (iii) the persistently infected culture contained a heterogeneous mixture of defective Sindbis virus RNAs which were not packaged into extracellular particles. To determine whether these intracellular RNAs could interfere with the replication of Sin-16, we analyzed cells that were cloned from the persistently infected culture. One clone (A3) synthesized a single defective viral RNA which was lost with continued passaging in culture. Infection of A3 cells with Sin-16 showed that the presence of the defective RNA greatly enhanced cell survival and led to enrichment of this RNA. In contrast, cured cells were highly susceptible to killing by Sin-16, and survivors did not synthesize this RNA. Thus, A3 cells were not genetically altered in their response to Sin-16, but were protected from the cytopathic effects of infection by an RNA with the characteristics of a defective-interfering RNA.

Animals↗
PubMed1982

The unstable wDZL mutation of Drosophila is caused by a 13 kilobase insertion that is imprecisely excised in phenotypic revertants.

We have analyzed the lesion in wDZL, a genetically unstable mutant allele of the eye color locus, white, of Drosophila melanogaster. We have cloned the DNA of the white locus region of flies carrying the wDZL allele and find a 13 kilobase insertion not present in the wild-type at the corresponding location. In 12 independent cases examined, reversion to a wild-type eye color phenotype correlates with the excision of a portion of this 13 kilobase insertion, indicating that the insertion is the cause of the mutation. The portion of the insertion that is excised in these eye color revertants is heterogeneous in size but appears to include the central 6 kilobases of the insertion in all cases. Many of these eye color revertants continue to undergo mutation at the white locus, indicating that the residual portion of the insertion in these revertants is sufficient to promote mutations.

Alleles↗
PubMed1982

FB elements are the common basis for the instability of the wDZL and wC Drosophila mutations.

The DNA insertions that cause the highly unstable mutations wC and wDZL share extensive homology with the FB family of transposable elements. FB elements carry long, internally repetitious, inverted terminal repeats and thus differ in structure from other transposable elements. Our results suggest that FB elements may excise and cause chromosomal rearrangements at unusually high frequencies. The wC insertion is a single FB element. The wDZL insertion differs in that it contains two FB elements, one at each terminus. The wC and wDZL insertions contain 4.0 and 6.5 kilobase nonhomologous segments between their terminal repeats. In contrast to the middle repetitive FB elements, the central segment of the wDZL insertion is single-copy and present at a fixed location in the wild-type genome. It has apparently been transposed by the action of flanking FB elements, causing the wDZL mutation at its new location.

Alleles↗
PubMed1982

Physical map of the white locus of Drosophila melanogaster.

The white locus of Drosophila melanogaster is a genetically well-characterized locus, mutations in which alter the degree of pattern of pigmentation of the eyes. Using a previously cloned DNA segment containing a portion of the white locus of a mutant allele, we have cloned and characterized the DNA of a 48-kilobase chromosomal region of the Canton S wild-type strain. We have mapped the positions, relative to restriction endonuclease cleavage sites, of several previously characterized chromosomal rearrangement breakpoints that bracket the while locus. These results define a segment of 14 kilobase that contains all of the white locus sequences necessary for the production of a wild-type eye color phenotype. By conventional criteria, no repetitive sequences are present within this 14-kilobase segment; however, we have identified an extremely weak DNA sequence homology between a portion of this segment and a chromosomal region in the vicinity of the zeste locus.

Animals↗
PubMed1981

The 5' termini of RNAs encoded by the transposable element copia.

The 5' termini of copia-specific RNAs in Drosophila melanogaster tissue culture cells were determined by S1 nuclease mapping and cap analysis. Both major copia RNAs share an identical set of heterogeneous 5' ends. Three major cap 1 structures M7GpppCmpUp, M7GpppCmpCp and M7GpppGmpUp together with several other minor caps were found. Almost all the 5' termini, as judged by S-1 nuclease mapping, were located either in a pyrimidine-rich part of the terminal direct repeat or apparently outside of the copia element, suggesting that a proportion of copia transcripts derive from promoters external to the genetic element.

Animals↗
PubMed1981

Cloning of DNA sequences from the white locus of D. melanogaster by a novel and general method.

We describe the isolation of a cloned DNA segment carrying unique sequences from the white locus of Drosophila melanogaster. Sequences within the cloned segment are shown to hybridize in situ to the white locus region on the polytene chromosomes of both wild-type strains and strains carrying chromosomal rearrangements whose breakpoints bracket the white locus. We further show that two small deficiency mutations, deleting white locus genetic elements but not those of complementation groups contiguous to white, delete the genomic sequences corresponding to a portion of the cloned segment. The strategy we have employed to isolate this cloned segment exploits the existence of an allele at the white locus containing a copy of a previously cloned transposable, reiterated DNA sequence element. We describe a simple, rapid method for retrieving cloned segments carrying a copy of the transposable element together with contiguous sequences corresponding to this allele. The strategy described is potentially general and we discuss its application to the cloning of the DNA sequences of other genes in Drosophila, including those identified only by genetic analysis and for which no RNA product is known.

Animals↗
PubMed1981

A DNA segment isolated from chromosomal site 67B in D. melanogaster contains four closely linked heat-shock genes.

cDNA clones coding for two different small heat-shock polypeptides were isolated. Both clones hybridize exclusively to the heat-shock puff site 67B, and restriction mapping of embryonic Drosophila melanogaster DNA showed that the two genes probably occur as single copies and are closely linked. The analysis was extended by isolating genomic clones, which contain these genes and two additional ones. The four different genes code for heat-induced poly(A)+ RNAs. These genes are clustered within an 11 kb segment and are separated by spacers of 1.0-4.7 kb. Three of the genes were found to exhibit alternating polarities. Thus in spite of their close linkage, the four heat-induced genes are most likely organized in individual transcription units.

Chromosomes↗
PubMed1980

Terminal repeats of the Drosophila transposable element copia: nucleotide sequence and genomic organization.

We have determined the nucleotide sequence of the terminal regions of two members of the copia sequence family of D. melanogaster. The first 276 bp at one end of a copia element are repeated in direct orientation at its other end. The direct repeats on a single copia element are identical to each other, but they differ by two nucleotide substitutions between the two elements which were examined; this suggests that during transposition only one direct repeat of the parent element is used as a template for both direct repeats of the transposed element. Each direct repeat itself contain a 17 bp imperfectly matched inveted terminal repetition. The ends of copia show significant sequence homology both to the yeast Ty1 element and to the integrated provirus of avian spleen necrosis virus, two other eucaryotic elements known to insert at many different chromosomal locations. Analysis of the genomic organization of the direct repeat sequence demonstrates that it seldom, if ever, occurs unlinked to an entire copia element.

Animals↗
PubMed1977

Coordinate regulation of protein synthesis and messenger RNA content during growth arrest of suspension Chinese hamster ovary cells.

We have found Chinese Hamster Ovary cells, cultured in suspension, are subject to growth control by serum. When suspended in medium containing 0.5% serum the cells becomes reversibly arrested in the beginning of the G1 phase of the cell cycle and can be maintained in this viable, nonproliferating state for several days. This system was used to examine the regulation of protein synthesis with growth rate. In particular, the experiments addressed the question whether mRNA content is the principal controlling factor determining the rate of protein synthesis. The rate of leucine incorporation in resting cells in low serum is 2-to 2.5-fold lower than that of cells growing in 10% serum. The steady-state number of cytoplasmic poly A (+) RNA molecules shows a proportional decrease, consistent with it being a determining factor controlling the rate of protein synthesis. Furthermore, the rate of production of poly A (+) and poly A (-) RNA appears to be regulated coordinately. Regulation of the rate of initiation of translation would result in fewer ribosomes bound per active message and/or a lower proportion of total mRNA's being active. Our measurements indicate that the fraction of cytoplasmic poly A (+) mRNA in polyribosomes and the relative degree of loading of each active poly A(+) mRNA with ribosomes is the same in resting and growing cells. Thus these cells resemble 3T6 and translational control does not appear to be an important part of the change in protein synthetic rate with teh state of growth.

Animals↗
PubMed1976

Changes in RNA in relation to growth of the fibroblast. IV. Alterations in theproduction and processing of mRNA and rRNA in resting and growing cells.

In previous reports, it was shown that both the concentration and rate of production of rRNA and mRNA were greater in growing than in resting 3T6 fibroblasts. Studies on isolated nuclei indicated that ribosomal RNA production is apparently controlled at the level of transcription. In contrast, hnRNA, the putative precursor of mRNA, appeared to be synthesized at the same rate in resting and growing cells. This finding was unexpected and has been tested in several ways. In this report, we show by an independent method that the relative rate of production of mRNA compared to hnRNA is several-fold higher in growing than in resting cells. However, the kinetics of processing of mRNA appear unchanged. This result suggests either that mRNA arises from a small subfraction of hnRNA or that the efficiency of processing of the hnRNA precursor is an important control mechanism which determines mRNA production ingrowing and resting states. Comparison of the initial rates of labeling of hnRNA and cytoplasmic message gives the efficiency with which the cytoplasmic mRNA is produced from nucleoplasmic RNA. The very low efficiency (3-4% in growing and 1-2% in resting cells) suggests that not every hnRNA molecule gives rise to a cytoplasmic message. In contrast to the similar kinetics of mRNA production in resting and growing states, processing of ribosomal RNA is much slower in the resting state and the emergence time for 28S RNA from nucleolus is greatly lengthened.

Cell Division↗