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M A Osley

Publications and source records attributed to M A Osley.

32 records · Page 2Linked to original sources
PubMed1987

Trans-acting regulatory mutations that alter transcription of Saccharomyces cerevisiae histone genes.

Using a Saccharomyces cerevisiae strain containing an integrated copy of an H2A-lacZ fusion gene, we screened for mutants which overexpressed beta-galactosidase as a way to identify genes which regulate transcription of the histone genes. Five recessive mutants with this phenotype were shown to contain altered regulatory genes because they had lost repression of HTA1 transcription which occurs upon inhibition of chromosome replication (D. E. Lycan, M. A. Osley, and L. Hereford, Mol. Cell. Biol. 7:614-621, 1987). Periodic transcription was affected in the mutants as well, since the HTA1 gene was transcribed during the G1 and G2 phases of the cell cycle, periods in the cell cycle when this gene is normally not expressed. A similar loss of cell cycle-dependent transcription was noted for two of the three remaining histone loci, while the HO and CDC9 genes continued to be expressed periodically. Using isolated promoter elements inserted into a heterologous cycl-lacZ fusion gene, we demonstrated that the mutations fell in genes which acted through a negative site in the TRT1 H2A-H2B promoter.

Cell Cycle↗
PubMed1987

Role of transcriptional and posttranscriptional regulation in expression of histone genes in Saccharomyces cerevisiae.

We analyzed the role of posttranscriptional mechanisms in the regulation of histone gene expression in Saccharomyces cerevisiae. The rapid drop in histone RNA levels associated with the inhibition of ongoing DNA replication was postulated to be due to posttranscriptional degradation of histone transcripts. However, in analyzing the sequences required for this response, we showed that the coupling of histone RNA levels to DNA replication was due mostly, if not entirely, to transcriptional regulatory mechanisms. Furthermore, deletions which removed the negative, cell cycle control sequences from the histone promoter also uncoupled histone transcription from DNA replication. We propose that the arrest of DNA synthesis prematurely activates the regulatory pathway used in the normal cell cycle to repress transcription. Although posttranscriptional regulation did not appear to play a significant role in coupling histone RNA levels to DNA replication, it did affect the levels of histone RNA in the cell cycle. Posttranscriptional regulation could apparently restore much of the periodicity of histone RNA accumulation in cells which constitutively transcribed the histone genes. Unlike transcriptional regulation, periodic posttranscriptional regulation appears to operate on a clock which is independent of events in the mitotic DNA cycle. Posttranscriptional recognition of histone RNA must require either sequences in the 3' end of the RNA or an intact three-dimensional structure since H2A- and H2B-lacZ fusion transcripts, containing only 5' histone sequences, were insensitive to posttranscriptional controls.

Cell Cycle↗
PubMed1986

Identification of sequences in a yeast histone promoter involved in periodic transcription.

Sequences between a pair of divergently transcribed histone genes in Saccharomyces cerevisiae are able to confer periodic transcription during the cell cycle. This conclusion contrasts to our previous hypothesis that an ars (autonomously replicating sequence) 3' to this locus is a transcription timer for yeast histone genes. The promoter sequences required for periodic expression have been localized by deletion analysis, and isolated elements have been analyzed by insertion into a heterologous promoter. Two cell-cycle-specific promoter functions have been identified. One function activates transcription in a cell-cycle-dependent manner. The other periodically represses transcription. Negative regulation may be the predominant form of cell-cycle control, because removal of the repressing function results in constitutive expression of the histone genes.

Base Sequence↗
PubMed1985

Regulation of CDC9, the Saccharomyces cerevisiae gene that encodes DNA ligase.

We have cloned CDC9, the structural gene for Saccharomyces cerevisiae DNA ligase, and investigated its transcriptional regulation both as a function of cell cycle stage and after UV irradiation. The steady-state level of DNA ligase mRNA increases at least fourfold in late G1, after the completion of start but before S phase. This high level of CDC9 mRNA then decays with an apparent half-life of ca. 20 min and remains at a low basal level throughout the rest of the cell cycle. The accumulation of CDC9 mRNA in late G1 is dependent upon the completion of start but not the CDC7 and CDC8 functions. Exposure of cells to UV light elicits an eightfold increase in DNA ligase mRNA levels.

Cell Cycle↗
PubMed1982

Periodic transcription of yeast histone genes.

Periodic transcription of yeast histone genes has been demonstrated by DNA excess filter hybridization of in vivo pulse-labeled RNA isolated from synchronous cell cultures. Using strains carrying cell division cycle (cdc) mutations, we show that both activation and termination of transcription are determined by temporally separable (cell cycle) events. Activation of histone mRNA synthesis occurs late in G1, at a point prior to initiation of DNA replication. Cessation of histone mRNA synthesis, however, is dependent upon the entry of cells into S. These results suggest a simple model for the control of histone gene transcription in which changes in chromatin that must precede the initiation of DNA replication also bring about activation of histone mRNA synthesis. Cessation of synthesis would occur once this region had been replicated and the chromatin restored to its prereplicative state.

DNA Replication↗
PubMed1982

Identification of a sequence responsible for periodic synthesis of yeast histone 2A mRNA.

Sequences required for the regulated expression of a yeast histone 2A (H2A) gene have been investigated by using fusions between this gene and the Escherichia coli lacZ gene. Fusions containing the entire spacer region in which divergent transcription of the H2A and H2B genes is initiated result in low-level constitutive synthesis of beta-galactosidase (beta-D-galactoside galactohydrolase, EC 3.2.1.23) in yeast. Regulated expression (which is characterized by periodic synthesis during the S phase of the cell cycle) is restored when a 1.3-kilobase HindIII fragment containing a small region of the 3' end of the H2B gene is present in either orientation. The regulatory activity in this region appears to be coincident with a sequence that supports autonomous replication in yeast.

Base Sequence↗
PubMed1982

Circular organization of the DNA synthetic pathway in Caulobacter crescentus.

Genetic analysis of the cell cycle of Caulobacter crescentus has identified a DNA synthetic pathway and a cell division pathway (M. A. Osley and A. Newton, J. Mol. Biol. 138:109-128, 1980). The results presented here show that in double-shift experiments the function of the PC2076 gene product, which is required for the initiation of DNA synthesis, depends on completion of a late stage of chromosome replication in the previous cell cycle. These findings suggest a circular organization of steps in the DNA synthetic pathway in C. crescentus.

Bacteria↗
PubMed1981

Cell-cycle regulation of yeast histone mRNA.

The levels of H2A and H2B mRNAs as a function of cell-cycle stage were determined by hybridization methods. The analysis was extended to H3 and H4 mRNAs by in vitro translation. Cells were partitioned into cell-cycle stages either by centrifugal elutriation or by G1 synchronization with the yeast mating pheromone, alpha factor. The data lead to the following conclusions. First, histone mRNA can be detected in significant quantities only in S-phase cells. Second, the point of maximal accumulation of histone mRNA is not coincident with the point of maximal DNA synthesis; rather, histone mRNA begins accumulating very early in S, reaching a maximum when less than one half of the DNA has replicated. From this point in the cell cycle the histone mRNA levels decrease, reaching basal levels at the end of S. Third, in spite of the fact that the rate of histone mRNA accumulation is not coincident with the rate of DNA synthesis, the two processes are coupled; inhibition of DNA synthesis results in an extremely rapid disappearance of histone mRNA that is much shorter than the normal histone mRNA half-life. Fourth, there is no visible accumulation of mRNA precursors at any cell-cycle stage. We can conclude that, in yeast, histone mRNA levels are tightly and coordinately regulated throughout cell division and that this regulation most likely occurs at both transcriptional and posttranscriptional levels. We also show that the two genetically unlinked H2B genes present in yeast are both expressed at comparable levels and are regulated. The regulation is probably sequence-specific, since genes in close proximity to the histones are not subject to cell-cycle control.

Cell Cycle↗
PubMed1981

Yeast histone genes show dosage compensation.

The copy number of yeast histone genes was increased by inserting an extra H2A,H2B gene pair into the haploid genome by the technique of yeast transformation. The presence of this extra gene copy has no detectable effect on cell growth. The steady-state levels of histone H2A,H2B mRNAs are not elevated in transformed strains, and they correspond to the levels measured for the parental strain. The transcription rate is increased in these strains, however, and the parental steady-state levels of histone mRNAs are maintained by increased turnover of histone transcripts. These results demonstrate that yeast histone genes display dosage compensation through the operation of posttranscriptional controls. They also suggest that maintainance of a constant ratio between histone mRNA concentration and the rate of chromosome replication may be of general importance to histone mRNA metabolism.

Gene Expression Regulation↗
PubMed1978

Regulation of cell cycle events in asymmetrically dividing cells: functions required for DNA initiation and chain elongation in Caulobacter crescentus.

To study the regulation of cell cycle events after asymmetric cell division in Caulobacter crescentus, we have identified functions that are required for DNA synthesis in the stalked cell produced at division and in the new stalked cell that develops from the swarmer cell 60 min after division. The initiation of DNA synthesis in the two progeny cells is dependent upon at least two common functions. One of these is a requirement for protein synthesis and the other is a gene product identified in a temperature-sensitive cell cycle mutant. DNA chain elongation requires a third common function. The characteristic pattern of DNA synthesis in C. crescentus appears to be controlled in part by the expression of these functions in the two stalked cells at different times after cell division. The age distribution for Caulobacter cells in an exponential population has been calculated (Appendix by Robert Tax) and used to analyze some of the results.

Bacteria↗
PubMed1977

Regulation of flagellin synthesis in the cell cycle of caulobacter: dependence on DNA replication.

Synthesis of the two filament proteins (flagellin A and flagellin B) of the Caulobacter creascentus flagellum was measured during the cell cycle. Synchronous cells were pulse-labeled with 36S-methionine, and flagellin proteins were isolated from crude extracts by radioimmune precipitation. The results showed that both proteins are maximally induced during the G2 period and that their induction requires de novo transcription. Flagellin A, however, continues to be made in the progeny swarmer cells after flagellin B synthesis has stopped. This discoordination in flagellin A and B synthesis and the relative abundance of the two proteins may result in part from the longer functional half-life of the flagellin A messenger RNA. Analysis of temperature-sensitive DNA chain elongation mutants suggests that the periodicity of flagellin A and B synthesis in the cell cycle is controlled by a late cell cycle event, presumably the completion of chromosome replication.

Bacteria↗
PubMed1977

Mutational analysis of developmental control in Caulobacter crescentus.

The relationship between the cell cycle and control of development has been studied by a genetic analysis of Caulobacter crescentus. The behavior of conditional cell division mutants showed that cell cycle events, such as DNA replication and cell division, are organized into a dependent pathway(s), i.e., later steps cannot preceed until earlier ones are completed. The ability of these strains to develop normally under nonpermissive conditions suggested that flagellin synthesis and stalk formation are dependent on the completion of different cell-cycle steps: the periodicity of flagellin synthesis is achieved by coupling it to either DNA chain elongation or completion, and stalk formation is dependent on a later step in the cell-division pathway. These developmental events are not themselves required for cell division, however.

Bacteria↗