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Mapping of a Physarum chromosomal origin of replication tightly linked to a developmentally-regulated profilin gene
1Laboratoire de Biologie et Ultrastructure du Noyau, UPR 272, CNRS, Villejuif, France.
Abstract:
We compared the pattern of replication of two cell-type specific profilin genes in one developmental stage of the slime mold Physarum polycephalum. Taking advantage of the natural synchrony of S-phase within the plasmodium, we established that the actively transcribed profilin P gene is tightly linked to a chromosomal replication origin and is replicated at the onset of S-phase. In contrast, the inactive profilin A gene is not associated with a replication origin and it is duplicated in mid S-phase. Mapping by two-dimensional gel electrophoresis defines a short DNA fragment in the proximal upstream region of the profilin P gene from which bidirectional replication is initiated. We further provide an estimate of the kinetics of elongation of the replicon and demonstrate that the 2 alleles of the profilin P gene are coordinately replicated. All these results were obtained on total DNA preparations extracted from untreated cells. They provide a strong evidence for site specific initiation of DNA replication in Physarum.
Insights
Replication timing of slime mold genes reveals distinct patterns. Actively transcribed profilin P genes initiate DNA replication early, while inactive profilin A genes replicate later in S-phase.
Area of Science:
- Molecular Biology
- Cell Biology
- Genetics
Background:
- Cell-type specific genes play crucial roles in development.
- Understanding gene replication timing is key to gene regulation.
- Physarum polycephalum offers a model for studying DNA replication due to its naturally synchronized S-phase.
Purpose of the Study:
- To compare the replication patterns of two cell-type specific profilin genes in Physarum polycephalum.
- To investigate the relationship between gene transcription and replication timing.
- To identify the specific DNA sequences involved in replication initiation.
Main Methods:
- Utilizing the naturally synchronized S-phase of Physarum polycephalum.
- Employing two-dimensional gel electrophoresis for DNA mapping.
- Analyzing total DNA preparations from untreated cells.
Main Results:
- The actively transcribed profilin P gene is linked to a replication origin and replicates at the start of S-phase.
- The inactive profilin A gene lacks an associated replication origin and replicates in mid S-phase.
- A specific upstream DNA fragment of the profilin P gene was identified as the replication initiation site, with bidirectional replication observed.
Conclusions:
- DNA replication initiation in Physarum is site-specific.
- Replication timing is correlated with gene activity and chromosomal organization.
- The findings provide insights into the regulation of gene replication during development.