Abstract
The RB/E2F pathway is involved in the control of the G(1)/S transition of the eukaryotic cell cycle where various S phase genes are activated by specific E2F factors. Ribonucleotide reductase (RNR) plays an essential role in the DNA synthesis pathway. Earlier studies showed that there are at least two RNR1 genes (RNR1a and RNR1b) and one RNR2 gene in tobacco. In synchronized tobacco BY2 cells, RNR1b gene expression is at its highest level in S phase. To investigate transcriptional regulation of the RNR1b gene, its promoter region was cloned and sequenced. Unlike its animal counterparts, the tobacco RNR1b promoter contains a consensus E2F-binding site. Surprisingly, this site is found in the leader sequence of the gene. We show here by gel shift analysis and antibody competition that one nuclear complex specifically binds this motif, and an E2F factor is part of this complex. Using reporter gene analysis, tobacco RNR1b promoter activity was detected during S phase in synchronized cells and in plant meristematic tissues. Mutation of the E2F element substantially reduced both activities. For the first time in plants, a single E2F motif found in the leader sequence plays an important role in the meristem and S phase-specific expression of the tobacco RNR1b gene.
Highlights
Progression of the cell cycle is associated with the phasespecific induction of genes whose products control the cell cycle or are involved in DNA replication [1]
The retinoblasma-type proteins (RB)/E2F pathway is involved in the control of the G1/S transition of the eukaryotic cell cycle where various S phase genes are activated by specific E2F factors
E2F sites are not involved in S phase-specific expression of Ribonucleotide reductase (RNR) genes in mammals: RNR cell cycle regulation is mediated by a TFII-I binding initiator element in the mouse RNR1 promoter [21], whereas a proximal NF-Y binding element is involved in S phase induction of mouse RNR2 gene [22]
Summary
Progression of the cell cycle is associated with the phasespecific induction of genes whose products control the cell cycle or are involved in DNA replication [1]. The RB/E2F pathway is involved in the control of the G1/S transition of the eukaryotic cell cycle where various S phase genes are activated by specific E2F factors. Tobacco RNR1b promoter activity was detected during S phase in synchronized cells and in plant meristematic tissues.
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