Gilbert Engler

cdc2a expression in Arabidopsis is linked with competence for cell division.

A key regulator of the cell cycle is a highly conserved protein kinase whose catalytic subunit, p34(cdc2), is encoded by the cdc2 gene. We studied the control of the expression of the Arabidopsis cdc2a gene in cell suspensions and during plant development. In cell cultures, arrest of the cell cycle did not significantly affect cdc2a mRNA levels, but nutrient conditions were important for cdc2a expression. During plant development, the pattern of cdc2a expression was strongly correlated with the cell proliferation potential. The effects of external signals on cdc2a expression were analyzed. Wounding induced expression in leaves. Lack of light altered temporal regulation of cdc2a in the apical but not root meristem of seedlings. Differential cdc2a responses were obtained after different hormone treatments. Signals present only in intact plants were necessary to mediate these responses. Although other control levels have yet to be analyzed, these results suggest that the regulation of cdc2a expression may contribute greatly to spatial and temporal regulation of cell division in plants. Our results also show that cdc2a expression is not always coupled with cell proliferation but always precedes it. We propose that cdc2a expression may reflect a state of competence to divide, and that the release of other controls is necessary for cell division to occur.

Control of proliferation, endoreduplication and differentiation by the Arabidopsis E2Fa-DPa transcription factor.

New plant cells arise at the meristems, where they divide a few times before they leave the cell‐cycle program and start to differentiate. Here we show that the E2Fa–DPa transcription factor of Arabidopsis thaliana is a key regulator determining the proliferative status of plant cells. Ectopic expression of E2Fa induced sustained cell proliferation in normally differentiated cotyledon and hypocotyl cells. The phenotype was enhanced strongly by the co‐expression of E2Fa with its dimerization partner, DPa. In endoreduplicating cells, E2Fa–DPa also caused extra DNA replication that was correlated with transcriptional induction of S phase genes. Because E2Fa–DPa transgenic plants arrested early in development, we argue that controlled exit of the cell cycle is a prerequisite for normal plant development.

The functional organization of the nopaline A. tumefaciens plasmid pTiC58

Abstract We have employed the P type plasmid RP4 and the transposons Tn1 and Tn7 to isolate insertion and deletion mutations in the nopaline Ti-plasmid pTiC58. Mutations that inactivate all known Ti phenotypes have been located on the physical map. Most importantly, we have positioned several regions involved in the determination of oncogenicity. They correspond to regions of homology between octopine and nopaline plasmids. One of these regions is part of the T-DNA, the Ti-plasmid DNA present in transformed plant cells. There are also segments of the T-DNA that are not essential for oncogenicity. One of these determines the biosynthesis of nopaline in tumors. The latter regions might allow insertion of foreign DNA that can then be introduced into plant cells.

A Mutation of the Mitochondrial ABC Transporter Sta1 Leads to Dwarfism and Chlorosis in the Arabidopsis Mutant starik

A mutation in the Arabidopsis gene STARIK leads to dwarfism and chlorosis of plants with an altered morphology of leaf and cell nuclei. We show that the STARIK gene encodes the mitochondrial ABC transporter Sta1 that belongs to a subfamily of Arabidopsis half-ABC transporters. The severity of the starik phenotype is suppressed by the ectopic expression of the STA2 homolog; thus, Sta1 function is partially redundant. Sta1 supports the maturation of cytosolic Fe/S protein in Δatm1 yeast, substituting for the ABC transporter Atm1p. Similar to Atm1p-deficient yeast, mitochondria of the starik mutant accumulated more nonheme, nonprotein iron than did wild-type organelles. We further show that plant mitochondria contain a putative l-cysteine desulfurase. Taken together, our results suggest that plant mitochondria possess an evolutionarily conserved Fe/S cluster biosynthesis pathway, which is linked to the intracellular iron homeostasis by the function of Atm1p-like ABC transporters.

Molecular Markers and Cell Cycle Inhibitors Show the Importance of Cell Cycle Progression in Nematode-Induced Galls and Syncytia

Root knot and cyst nematodes induce large multinucleated cells, designated giant cells and syncytia, respectively, in plant roots. We have used molecular markers to study cell cycle progression in these specialized feeding cells. In situ hybridization with two cyclin-dependent kinases and two cyclins showed that these genes were induced very early in galls and syncytia and that the feeding cells progressed through the G2 phase. By using cell cycle blockers, DNA synthesis and progression through the G2 phase, or mitosis, were shown to be essential for gall and syncytium establishment. When mitosis was blocked, further gall development was arrested. This result demonstrates that cycles of endoreduplication or other methods of DNA amplification are insufficient to drive giant cell expansion. On the other hand, syncytium development was much less affected by a mitotic block; however, syncytium expansion was inhibited.

PtABI3 Impinges on the Growth and Differentiation of Embryonic Leaves during Bud Set in Poplar

The Arabidopsis ABSCISIC ACID–INSENSITIVE3 (ABI3) protein plays a crucial role during late seed development and has an additional function at the vegetative meristem, particularly during periods of growth-arresting conditions and quiescence. Here, we show that the ABI3 homolog of poplar (PtABI3) is expressed in buds during natural bud set. Expression occurs clearly after perception of the critical daylength that initiates bud set and dormancy in poplar. In short-day conditions mimicking natural bud set, the expression of a chimeric PtABI3::β-glucuronidase (GUS) gene occurred in those organs and cells of the apex that grow actively but will undergo arrest: the young embryonic leaves, the subapical meristem, and the procambial strands. If PtABI3 is overexpressed or downregulated, bud development in short-day conditions is altered. Constitutive overexpression of PtABI3 resulted in apical buds with large embryonic leaves and small stipules, whereas in antisense lines, bud scales were large and leaves were small. Thus, PtABI3 influences the size and ratio of embryonic leaves and bud scales/stipules that differentiate from the primordia under short-day conditions. These observations, together with the expression of PtABI3::GUS in embryonic leaves but not in bud scales/stipules, support the idea that wild-type PtABI3 is required for the relative growth rate and differentiation of embryonic leaves inside the bud. These experiments reveal that ABI3 plays a role in the cellular differentiation of vegetative tissues, in addition to its function in seeds.

Transcriptome analysis during cell division in plants

Using synchronized tobacco Bright Yellow-2 cells and cDNA-amplified fragment length polymorphism-based genomewide expression analysis, we built a comprehensive collection of plant cell cycle-modulated genes. Approximately 1,340 periodically expressed genes were identified, including known cell cycle control genes as well as numerous unique candidate regulatory genes. A number of plant-specific genes were found to be cell cycle modulated. Other transcript tags were derived from unknown plant genes showing homology to cell cycle-regulatory genes of other organisms. Many of the genes encode novel or uncharacterized proteins, indicating that several processes underlying cell division are still largely unknown.

The Arabidopsis 1-Aminocyclopropane-1-Carboxylate Synthase Gene 1 Is Expressed during Early Development.

The temporal and spatial expression of one member of the Arabidopsis 1-aminocyclopropane-1-carboxylate (ACC) synthase gene family (ACS1) was analyzed using a promoter-[beta]-glucuronidase fusion. The expression of ACS1 is under developmental control both in shoot and root. High expression was observed in young tissues and was switched off in mature tissues. ACS1 promoter activity was strongly correlated with lateral root formation. Dark-grown seedlings exhibited a different expression pattern from light-grown ones. The ACC content and the in vivo activity of ACC oxidase were determined. ACC content correlated with ACS1 gene activity. ACC oxidase activity was demonstrated in young Arabidopsis seedlings. Thus, the ACC formed can be converted into ethylene. In addition, ethylene production of immature leaves was fourfold higher compared to that of mature leaves. The possible involvement of ACS1 in influencing plant growth and development is discussed.

A new D-type cyclin of Arabidopsis thaliana expressed during lateral root primordia formation

Abstract. D-type cyclins are believed to regulate the onset of cell division upon mitogenic signaling. Here, the isolation is reported of a new D-type cyclin gene (CYCD4;1) of Arabidopsis thaliana (L.) Heynh. during a two-hybrid screen using the cyclin-dependent kinase CDC2aAt as bait. Transcription of CYCD4;1 can be induced by sucrose. The co-regulated expression of CYCD4;1 and CDC2aAt in starved suspension cultures upon mitogenic stimulation indicates that the formation of a complex between these two partners is important for the resumption of cell division activity. By in-situ hybridizations CYCD4;1 was shown to be expressed during vascular tissue development, embryogenesis, and formation of lateral root primordia. Expression during the latter process suggests that the induced expression of D-type cyclins by mitogenic stimuli might be one of the rate-limiting events for the initiation of lateral roots.

An easy technique for the clearing of histochemically stained plant tissue

The sites of GUS-activity in strongly pigmented whole-mount preparations or thick handmade sections of plant material can easily be visualized by clearing it with chlorallactophenol (CLP), without affecting the ClBr-indigo precipitate.