Alfalfa Cyclins: Differential Expression during the Cell Cycle and in Plant Organs

Ran is a 25-kDa Ras-related nuclear GTPbinding protein which is very highly conserved in humans, Saccharomyces cerevisiae, and Schizosaccharomyces pombe. Ran has been found to form a stable, noncovalent complex with the chromatin-associated protein RCC1, a negative regulator of mitosis. In Sch. pombe, a temperature-sensitive mutation in the RCC1 homolog encoded by thepiml gene causes premature induction of mitosis, and this mutation can be suppressed by overexpression of the Ran homolog encoded by spil. We report here the cloning of three Ran cDNAs from tomato. The Ras-related protein Ran is a 25-kDa nuclear protein that was shown in mammalian cells to bind to the chromatinassociated protein RCC1 (1). RCC1 was originally isolated from a hamster cell line (2) where a temperature-sensitive RCC1 mutation causes either premature chromosome condensation, activation ofp34cdc2 kinase, and entry into mitosis if the temperature is shifted during the S or G2 phase of the cell cycle or cell-cycle arrest if the temperature is shifted during G1 (3, 4). RCC1 catalyzes the exchange of guanine nucleotides on the Ran protein (5), suggesting that RCC1 and Ran may constitute parts of a GTPase switch that is involved in the coupling of the completion of DNA replication to the onset of M phase (for reviews, see refs. 6-8). Like their mammalian counterparts, the RCC1 and Ran homologs of Schizosaccharomyces pombe, called pimil and spil, respectively, have also been shown to be involved in the coupling of DNA replication to mitosis (9). A temperature-sensitive mutation in the piml gene causes premature chromosome condensation and entry into mitosis at the restrictive temperature. Overexpression of spil can suppress this mutation, and piml and spil have been shown to physically interact in vitro (10). In Saccharomyces cerevisiae a temperaturesensitive mutation in the RCC1 homolog PRP20/SRM1 can be suppressed by overexpression of either of the two S. cerevisiae Ran homologs (11,12). However, unlike RCC1 and piml, PRP20/SRM1 was first discovered as a gene essential for a variety of nuclear functions including pheromone response, mRNA metabolism and export, nuclear structure, and plasmid and chromosome stability (13-17). Also, H1 kinase is not activated in srml mutants (16), and disruption of the two Ran genes inhibits mRNA export (12), indicating that RCC1 and Ran play broad roles in nuclear structure and function in S. cerevisiae, which may differ from other organisms. However, findings that RCC1 is involved in mRNA export from mammalian nuclei (14) (22). Their sequences were 5'-GGGAATTCGGIGARTTYGAGAARAARTA-3' and 5'-CGGGATCCGGYTTYTCGAARTTRTARTT-3', where I is deoxyinosine. The PCR mixture contained 500 pmol of each primer, amplified phage library from tomato (1.2 x 107 plaque-forming units) or Arabidopsis (1.7 x 106 plaqueforming units), and 2.5 units of Taq DNA polymerase (Perkin-Elmer/Cetus). Forty-four cycles were performed of 1 min at 94°C, 2 min at the annealing temperature, and 2 min at 70°C. The annealing temperature star...

Section: Akqvtfhrkknlqyyeisaksnynfekpflylarklagdgnlhfv Tomato 2 -----mentioning

confidence: 81%

A small nuclear GTP-binding protein from tomato suppresses a Schizosaccharomyces pombe cell-cycle mutant.

Ach

Gruissem²

1994

“…Homologs ofthe cdc2 gene have been found in plants (10)(11)(12)(13)(14), and two putative mitotic cyclins have been cloned, from soybean (15) and from Arabidopsis (16). Partial cDNA clones believed to encode cyclins have also been isolated from carrot (15) and alfalfa (17). In this report, we show that several putative mitotic cyclins capable of inducing maturation in Xenopus oocytes are present in the same plant species (maize).…”

mentioning

confidence: 79%

“…In addition, cyclin homologous cDNAs have been isolated from carrot (15) and alfalfa (17), but the putative cyclins encoded by them have lower homologies to the cyclins from soybean and Arabidopsis and to each other (15,17). It is not known whether the partial cDNA clones from carrot (15) and alfalfa (17) represent functional homologs of mitotic cyclins and, if so, whether they represent species differences or different structural groups of plant cyclins.…”

Section: Discussionmentioning

confidence: 99%

Cloning of four cyclins from maize indicates that higher plants have three structurally distinct groups of mitotic cyclins.

Renaudin

Colasanti

Rime

et al. 1994

100

While a large number of cydins have been described in animals and yeasts, very limited information is available regarding cydins in plants. We describe here the isolation of cDNA clones encoding four putative mitotic cyclins from maize. AU four cyclins were able to induce maturation of Xenopus oocytes, demonstrating that they can act as mitotic cyclins in this system. Northern analysis showed that all four cyclins were expressed only in actively dividing tissues and organs, with a stronger correlation between expression and mitotic activity than is observed with cdc2. The deduced protein sequences suggest that the four maize cyclins belong to the cyclin A and B families identified from animal and yeast studies but that they cannot be described easily as either A-type or B-type cyclins. However, comparison with previously cloned plant cyclins shows that cyclins in higher plants form three distinct structural groups that have been conserved in both monocotyledonous and dicotyledonous species and that cyclins from all three groups are present within a single plant species.Studies on the mechanism of cell division control in eukaryotic cells (1) have shown that protein kinases encoded by homologs of the Schizosaccharomyces pombe cdc2 gene (2), in association with proteins known as cyclins (3), play a key role in driving the cell cycle. Cyclins from the six structural types so far identified are presumed to form part of a kinase complex in association with p34cdc2 or closely related protein kinases of the cdk family (2, 4, 5). The large diversity of cyclins, in addition to that of their cdk counterparts, is believed to account for the specific phosphorylation of different sets of substrates at successive transitions of the cell cycle (2, 5).Significant differences exist between plants and animals in the regulation of cell division during development (6-9), but much less is known about cell cycle regulators in plants (8,9

“…The data presented here indicate that the cyc3aAt and cyclAt genes are expressed at a cell cycle window that resemble that of the A and B classes of animal cyclins, respectively. Studies in synchronized alfalfa (Medicago sativa) cells, and double-target in situ hybridization of Antirrhinum and soybean (Glycine max) plants, demonstrated that mRNA levels of the alfalfa cycMs2 gene, the Antirrhinum cyclAm and cyc2Am genes, and the soybean cyc5Gm gene, all more homologous to B cyclins, are abundant in G2 and M phase cells (28)(29)(30). Using doubletarget in situ hybridization, it was recently concluded that mRNA of the soybean cyc3Gm gene, which is more related to animal A cyclins, is expressed in late S phase and G2 cells (30).…”

Section: Discussionmentioning

confidence: 99%

Two Arabidopsis cyclin promoters mediate distinctive transcriptional oscillation in synchronized tobacco BY-2 cells.

Shaul

Mironov

Burssens

et al. 1996

169

110

Cyclins are cell cycle regulators whose proteins oscillate dramatically during the cell cycle. Cyclin steady-state mRNA levels also fluctuate, and there are indications that both their rate of transcription and mRNA stability are under cell cycle control. Here, we demonstrate the transcriptional regulation of higher eukaryote cyclins throughout the whole cell cycle with a high temporal resolution. The promoters of two Arabidopsis cyclins, cyc3aAt and cyclAt, mediated transcriptional oscillation of the 8-glucuronidase (gus) reporter gene in stably transformed tobacco BY-2 cell lines. The rate of transcription driven by the cyc3aAt promoter was very low during G1, slowly increased during the S phase, peaked at the G2 phase and G2-to-M transition, and was down-regulated before early metaphase. In contrast, the rate of the cyclAt-related transcription increased upon exit of the S phase, peaked at the G2-to-M transition and during mitosis, and decreased upon exit from the M phase. This study indicates that transcription mechanisms that seem to be conserved among species play a significant role in regulating the mRNA abundance of the plant cyclins. Furthermore, the transcription patterns of cyc3aAt and cyclAt were coherent with their slightly higher sequence similarity to the A and B groups of animal cyclins, respectively, suggesting that they may fulfill comparable roles during the cell cycle.Cyclins are activators of specific serine/threonine protein kinases, termed CDKs, which drive progression of the eukaryotic cell cycle (reviewed in ref. 1). Based upon sequence analyses, most plant cyclins identified so far can be divided into those showing slightly higher sequence similarity to either the A or B groups of animal cyclins, but these similarities are not sufficient to exclusively assign them to either group (2, 3). Animal A-and B-type cyclins have distinct patterns of expression and fulfill different roles throughout the cell cycle (reviewed in ref. 4). As the roles of cyclins are far more understood in animals than in plants, further affiliation of a plant cyclin to a certain group of animal cyclins based on a similar expression pattern may give a clue to its function. The Arabidopsis cyclAt and cyc3aAt cyclin genes represent plant cyclins with slightly higher homology to the B and A groups of animal cyclins, respectively (2). Whole-mount in situ hybridization of Arabidopsis root tips treated with cell cycle blockers indicated that steady-state mRNA levels of cyclAt are high at early metaphase and low at early S phase, while the opposite was true for cyc3aAt (2). These data indicated that mRNA levels of cyc3aAt are increased in advance to that of cyclAt, but were not sufficient to give a complete picture of the expression pattern and transcriptional regulation of these cyclins throughout the whole cell cycle.Fluctuation in the steady-state mRNA level of a cell cycle gene may be regulated by a change in the rate of transcription, or in mRNA stability, or both. Transcriptional regulation of cell cycle genes...