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Root development is plastic, with post-embryonic organogenesis being mediated by meristems. Although cell division is intrinsic to meristem initiation, maintenance and proliferative growth, the role of the cell cycle in regulating growth and development is unclear. To address this question, we examined the expression of cdc2 and cye genes, which encode the catalytic and regulatory subunits, respectively, of cyclin-dependent protein kinases that control progression through the cell cycle. Unlike cdc2, which is expressed not only in apical meristems but also before lateral root initiation in quiescent, pericycle cells arrested in the G2 phase of the cell cycle, cyc1At transcripts accumulate specifically in dividing cells immediately before cytokinesis. Ectopic expression of cyc1At under the control of the cdc2aAt promoter in Arabidopsis plants markedly accelerates growth without altering the pattern of lateral root development or inducing neoplasia. Thus cyclin expression is a limiting factor for growth, which in turn drives indeterminate development of the root system.

The p34 protein kinase encoded by the cdc2 gene is a key component of the eukaryotic cell cycle required for the G1-to S-phase transition and entry into mitosis. To study the regulation of plant meristem activity and cell proliferation, we have examined the tissue-specific accumulation of cdc2 transcripts in Arabidopsis thlana and the related crucifer radish (Raphanus saivus) by in situ hybridization usingA. thaliana cdc2 cDNA sequences as a probe. cdc2 transcripts accumulated in leaf primordia and within the vegetative shoot apical meristem. During flower development, high levels of expression were observed in meristems, in the basal regions ofdeveloping organs, in the developing vasculature, and assocated with rib meristems elaborated late in the development ofsome floral organs. In root tips, cdc2 transcripts accumulated in the meristematic region and adjacent daughter cells but were not detected in the quiescent center. There was strong hybridization throughout the pericycle, and a further localized accumulation of cdc2 transcripts was observed in the initial stages ofthe activation ofa new meristem at sites of lateral root development. We conclude that cdc2 expression is a critical factor in the regulation of meristem activity and establishment of proliferative competence.The p34 protein kinase encoded by cdc2 is a key component of the eukaryotic cell cycle required for the Gj-to S-phase transition and entry into mitosis (1-3). p34 protein kinases become active only after forming a complex with a cyclin, and mutant analysis in Saccharomyces cerevisiae and Schizosaccharomyces pombe have demonstrated the presence of both positive regulators (e.g., cdc25+ and nim1 +) and negative regulators (e.g., wee1+) of M-phase p34 protein kinase activity (1-3). p34 protein kinases are highly conserved in evolution and have been found in all eukaryotes analyzed, including several plant species. cdc2 cDNA clones from Arabidopsis thaliana (4, 5), alfalfa (6), pea (7), and maize (8) complement cdc2 mutants in Sc. pombe or mutants in the equivalent gene (cdc28) in S. cerevisiae.Molecular cloning of plant cdc2 sequences provides the basis for mechanistic analysis of the regulation of cell proliferation in relation to the unique stem cell organization of higher plants mediating continuing organogenesis during postembryonic growth. In the present paper, we demonstrate by in situ hybridization analysis of the tissue-specific accumulation of cdc2 transcripts in A. thaliana and the related crucifer radish (Raphanus sativus) that the spatial and temporal pattern of cdc2 expression is a critical factor in regulation of meristem activity.MATERIALS AND METHODS Plant Material. A. thaliana ecotypes Columbia and Landsberg erecta were grown in the greenhouse under a 16-h light/8-h dark regimen. To isolate RNA from roots, plants were grown in liquid MS medium (9). For auxin induction, radish (cv. Scarlet Globe) seeds were surface sterilized and germinated in the dark (10). Roots were cut into 2-cm pieces and incubated in the dark at ...

To facilitate molecular studies of symbiotic nitrogen fixation a procedure for rapid production of transgenic root nodules was established on the legumeLotus corniculatus (Bird'sfoot trefoil). Regeneration of transgenic plants is not required as transgenic nodules are formed onAgrobacterium rhizogenes incited roots inoculated withRhizobium. Easy identification of transformed roots is possible using a set ofA. rhizogenes acceptor strains carrying assayable marker genes such as chloramphenicol acetyltransferase (CAT), β-glucuronidase (GUS), or luciferase (LUC) under control of the cauliflower mosaic virus (CaMV) 35S promoter. Counterselection ofA. rhizogenes after infection of plants was improved using an auxotrophy marker.