Transverse sectioning of Arabidopsis thaliana leaves using resin embedding

The leaf is the major functional part of the shoot performing the bulk of photosynthetic activity. Its development is relatively plastic allowing the plant to adapt to environmental changes by modifying leaf size and anatomy. Moreover, a leaf is made up of various distinct cell layers, each having specialized functions. To understand functional adaptation and the development of the leaf it is essential to obtain cross sections throughout leaf development and at maturity (Kalve et al., 2014). Here, we describe a protocol for transverse sectioning of Arabidopsis thaliana leaves using resin embedding. This protocol provides a reliable platform to yield high quality images of cross sections allowing study of development of various tissue layers across the transversal axis of the leaf. As this method is an adaptation of the protocol developed for the Arabidopsis root tip by Beeckman and Viane (1999) and De Smet et al. (2004), it can easily be modified to accommodate other organs and species

Numerical bifurcation analysis of pattern formation in a cell based auxin transport model

Transport models of growth hormones can be used to reproduce the hormone accumulations that occur in plant organs. Mostly, these accumulation patterns are calculated using time step methods, even though only the resulting steady state patterns of the model are of interest. We examine the steady state solutions of the hormone transport model of Smith et al (2006) for a one-dimensional row of plant cells. We search for the steady state solutions as a function of three of the model parameters by using numerical continuation methods and bifurcation analysis. These methods are more adequate for solving steady state problems than time step methods. We discuss a trivial solution where the concentrations of hormones are equal in all cells and examine its stability region. We identify two generic bifurcation scenarios through which the trivial solution loses its stability. The trivial solution becomes either a steady state pattern with regular spaced peaks or a pattern where the concentration is periodic in time.Comment: submitte

The plant-specific cyclin-dependent kinase CDKB1;1 and transcription factor E2Fa-DPa control the balance of mitotically dividing and endoreduplicating cells in Arabidopsis

Transgenic Arabidopsis thaliana plants overproducing the E2Fa-DPa transcription factor have two distinct cell-specific phenotypes: some cells divide ectopically and others are stimulated to endocycle. The decision of cells to undergo extra mitotic divisions has been postulated to depend on the presence of a mitosis-inducing factor (MIF). Plants possess a unique class of cyclin-dependent kinases (CDKs; B-type) for which no ortholog is found in other kingdoms. The peak of CDKB1;1 activity around the G2-M boundary suggested that it might be part of the MIF. Plants that overexpressed a dominant negative allele of CDKB1;1 underwent enhanced endoreduplication, demonstrating that CDKB1;1 activity was required to inhibit the endocycle. Moreover, when the mutant CDKB1;1 allele was overexpressed in an E2Fa-DPa–overproducing background, it enhanced the endoreduplication phenotype, whereas the extra mitotic cell divisions normally induced by E2Fa-DPa were repressed. Surprisingly, CDKB1;1 transcription was controlled by the E2F pathway, as shown by its upregulation in E2Fa-DPa–overproducing plants and mutational analysis of the E2F binding site in the CDKB1;1 promoter. These findings illustrate a cross talking mechanism between the G1-S and G2-M transition points

How grass keeps growing : an integrated analysis of hormonal crosstalk in the maize leaf growth zone

We studied the maize leaf to understand how long-distance signals, auxin and cytokinin, control leaf growth dynamics. We constructed a mathematical model describing the transport of these hormones along the leaf growth zone and their interaction with the local gibberellin (GA) metabolism in the control of cell division. Assuming gradually declining auxin and cytokinin supply at the leaf base, the model generated spatiotemporal hormone distribution and growth patterns that matched experimental data. At the cellular level, the model predicted a basal leaf growth as a result of cell division driven by auxin and cytokinin. Superimposed on this, GA synthesis regulated growth through the control of the size of the region of active cell division. The predicted hormone and cell length distributions closely matched experimental data. To correctly predict the leaf growth profiles and final organ size of lines with reduced or elevated GA production, the model required a signal proportional to the size of the emerged part of the leaf that inhibited the basal leaf growth driven by auxin and cytokinin. Excision and shading of the emerged part of the growing leaf allowed us to demonstrate that this signal exists and depends on the perception of light intensity

The cyclin-dependent kinase inhibitor Orysa;KRP1 plays an important role in seed development of rice

Kip-related proteins (KRPs) play a major role in the regulation of the plant cell cycle. We report the identification of five putative rice (Oryza sativa) proteins that share characteristic motifs with previously described plant KRPs. To investigate the function of KRPs in rice development, we generated transgenic plants overexpressing the Orysa; KRP1 gene. Phenotypic analysis revealed that overexpressed KRP1 reduced cell production during leaf development. The reduced cell production in the leaf meristem was partly compensated by an increased cell size, demonstrating the existence of a compensatory mechanism in monocot species by which growth rate is less reduced than cell production, through cell expansion. Furthermore, Orysa; KRP1 overexpression dramatically reduced seed filling. Sectioning through the overexpressed KRP1 seeds showed that KRP overproduction disturbed the production of endosperm cells. The decrease in the number of fully formed seeds was accompanied by a drop in the endoreduplication of endosperm cells, pointing toward a role of KRP1 in connecting endocycle with endosperm development. Also, spatial and temporal transcript detection in developing seeds suggests that Orysa; KRP1 plays an important role in the exit from the mitotic cell cycle during rice grain formation

Identification of putative cancer genes through data integration and comparative genomics between plants and humans

Coordination of cell division with growth and development is essential for the survival of organisms. Mistakes made during replication of genetic material can result in cell death, growth defects, or cancer. Because of the essential role of the molecular machinery that controls DNA replication and mitosis during development, its high degree of conservation among organisms is not surprising. Mammalian cell cycle genes have orthologues in plants, and vice versa. However, besides the many known and characterized proliferation genes, still undiscovered regulatory genes are expected to exist with conserved functions in plants and humans. Starting from genome-wide Arabidopsis thaliana microarray data, an integrative strategy based on coexpression, functional enrichment analysis, and cis-regulatory element annotation was combined with a comparative genomics approach between plants and humans to detect conserved cell cycle genes involved in DNA replication and/or DNA repair. With this systemic strategy, a set of 339 genes was identified as potentially conserved proliferation genes. Experimental analysis confirmed that 20 out of 40 selected genes had an impact on plant cell proliferation; likewise, an evolutionarily conserved role in cell division was corroborated for two human orthologues. Moreover, association analysis integrating Homo sapiens gene expression data with clinical information revealed that, for 45 genes, altered transcript levels and relapse risk clearly correlated. Our results illustrate how a systematic exploration of the A. thaliana genome can contribute to the experimental identification of new cell cycle regulators that might represent novel oncogenes or/and tumor suppressors

Drought induces distinct growth response, protection, and recovery mechanisms in the maize leaf growth zone

Drought is the most important crop yield-limiting factor, and detailed knowledge of its impact on plant growth regulation is crucial. The maize (Zea mays) leaf growth zone offers unique possibilities for studying the spatiotemporal regulation of developmental processes by transcriptional analyses and methods that require more material, such as metabolite and enzyme activity measurements. By means of a kinematic analysis, we show that drought inhibits maize leaf growth by inhibiting cell division in the meristem and cell expansion in the elongation zone. Through a microarray study, we observed the down- regulation of 32 of the 54 cell cycle genes, providing a basis for the inhibited cell division. We also found evidence for an up- regulation of the photosynthetic machinery and the antioxidant and redox systems. This was confirmed by increased chlorophyll content in mature cells and increased activity of antioxidant enzymes and metabolite levels across the growth zone, respectively. We demonstrate the functional significance of the identified transcriptional reprogramming by showing that increasing the antioxidant capacity in the proliferation zone, by overexpression of the Arabidopsis (Arabidopsis thaliana) iron-superoxide dismutase gene, increases leaf growth rate by stimulating cell division. We also show that the increased photosynthetic capacity leads to enhanced photosynthesis upon rewatering, facilitating the often-observed growth compensation.Fil: Avramova, Viktoriya. Universiteit Antwerp; BélgicaFil: Abdelgawad, Hamada. University of Beni-Suef; EgiptoFil: Zhang, Zhengfeng. Central China Normal University; ChinaFil: Fotschki, Bartosz. Institute of Animal Reproduction and Food Research; PoloniaFil: Casadevall, Romina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Centro de Estudios Fotosintéticos y Bioquímicos. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Centro de Estudios Fotosintéticos y Bioquímicos; ArgentinaFil: Vergauwen, Lucia. Universiteit Antwerp; BélgicaFil: Knapen, Dries. Universiteit Antwerp; BélgicaFil: Taleisnik, Edith. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigaciones Agropecuarias; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Guisez, Yves. Universiteit Antwerp; BélgicaFil: Asard, Han. Universiteit Antwerp; BélgicaFil: Beemster, Gerrit T. S.. Universiteit Antwerp; Bélgic