MDF is a conserved splicing factor and modulates cell division and stress response in .

The coordination of cell division with stress response is essential for maintaining genome stability in plant meristems. Proteins involved in pre-mRNA splicing are important for these processes in animal and human cells. Based on its homology to the splicing factor SART1, which is implicated in the control of cell division and genome stability in human cells, we analyzed if MDF has similar functions in plants.

γ-Tubulin Complexes and Fibrillar Arrays: Two Conserved High Molecular Forms with Many Cellular Functions.

Higher plants represent a large group of eukaryotes where centrosomes are absent. The functions of γ-tubulin small complexes (γ-TuSCs) and γ-tubulin ring complexes (γ-TuRCs) in metazoans and fungi in microtubule nucleation are well established and the majority of components found in the complexes are present in plants. However, plant microtubules are also nucleated in a γ-tubulin-dependent but γ-TuRC-independent manner.

γ-Tubulin interacts with E2F transcription factors to regulate proliferation and endocycling in Arabidopsis.

γ-Tubulin is associated with microtubule nucleation, but evidence is accumulating in eukaryotes that it also functions in nuclear processes and in cell division control independently of its canonical role. We found that in Arabidopsis thaliana, γ-tubulin interacts specifically with E2FA, E2FB, and E2FC transcription factors both in vitro and in vivo. The interaction of γ-tubulin with the E2Fs is not reduced in the presence of their dimerization partners (DPs) and, in agreement, we found that γ-tubulin interaction with E2Fs does not require the dimerization domain.

Tubulin: Structure, Functions and Roles in Disease.

Highly conserved α- and β-tubulin heterodimers assemble into dynamic microtubules and perform multiple important cellular functions such as structural support, pathway for transport and force generation in cell division. Tubulin exists in different forms of isotypes expressed by specific genes with spatially- and temporally-regulated expression levels. Some tubulin isotypes are differentially expressed in normal and neoplastic cells, providing a basis for cancer chemotherapy drug development.

Microtubular and Nuclear Functions of γ-Tubulin: Are They LINCed?

γ-Tubulin is a conserved member of the tubulin superfamily with a function in microtubule nucleation. Proteins of γ-tubulin complexes serve as nucleation templates as well as a majority of other proteins contributing to centrosomal and non-centrosomal nucleation, conserved across eukaryotes. There is a growing amount of evidence of γ-tubulin functions besides microtubule nucleation in transcription, DNA damage response, chromatin remodeling, and on its interactions with tumor suppressors.

γ-Tubulin has a conserved intrinsic property of self-polymerization into double stranded filaments and fibrillar networks.

γ-Tubulin is essential for microtubule nucleation and also plays less understood roles in nuclear and cell-cycle-related functions. High abundancy of γ-tubulin in acentrosomal Arabidopsis cells facilitated purification and biochemical characterization of large molecular species of γ-tubulin. TEM, fluorescence, and atomic force microscopy of purified high molecular γ-tubulin forms revealed the presence of linear filaments with a double protofilament substructure, filament bundles and aggregates.

RETINOBLASTOMA RELATED directly regulates DNA damage responses through functions beyond cell cycle control.

The rapidly proliferating cells in plant meristems must be protected from genome damage. Here, we show that the regulatory role of the RETINOBLASTOMA RELATED (RBR) in cell proliferation can be separated from a novel function in safeguarding genome integrity. Upon DNA damage, RBR and its binding partner E2FA are recruited to heterochromatic γH2AX-labelled DNA damage foci in an ATM- and ATR-dependent manner.

The Arabidopsis mitogen-activated protein kinase 6 is associated with γ-tubulin on microtubules, phosphorylates EB1c and maintains spindle orientation under nitrosative stress.

Stress-activated plant mitogen-activated protein (MAP) kinase pathways play roles in growth adaptation to the environment by modulating cell division through cytoskeletal regulation, but the mechanisms are poorly understood. We performed protein interaction and phosphorylation experiments with cytoskeletal proteins, mass spectrometric identification of MPK6 complexes and immunofluorescence analyses of the microtubular cytoskeleton of mitotic cells using wild-type, mpk6-2 mutant and plants overexpressing the MAP kinase-inactivating phosphatase, AP2C3. We showed that MPK6 interacted with γ-tubulin and co-sedimented with plant microtubules polymerized in vitro.

Overexpressed TPX2 causes ectopic formation of microtubular arrays in the nuclei of acentrosomal plant cells.

TPX2 performs multiple roles in microtubule organization. Previously, it was shown that plant AtTPX2 binds AtAurora1 kinase and colocalizes with microtubules in a cell cycle-specific manner. To elucidate the function of TPX2 further, this work analysed Arabidopsis cells overexpressing AtTPX2-GFP.

NITRILASE1 regulates the exit from proliferation, genome stability and plant development.

Nitrilases are highly conserved proteins with catabolic activity but much less understood functions in cell division and apoptosis. To elucidate the biological functions of Arabidopsis NITRILASE1, we characterized its molecular forms, cellular localization and involvement in cell proliferation and plant development. We performed biochemical and mass spectrometry analyses of NITRILASE1 complexes, electron microscopy of nitrilase polymers, imaging of developmental and cellular distribution, silencing and overexpression of nitrilases to study their functions.

Interactions of an Arabidopsis RanBPM homologue with LisH-CTLH domain proteins revealed high conservation of CTLH complexes in eukaryotes.

Background: RanBPM (Ran-binding protein in the microtubule-organizing centre) was originally reported as a centrosome-associated protein in human cells. However, RanBPM protein containing highly conserved SPRY, LisH, CTLH and CRA domains is currently considered as a scaffolding protein with multiple cellular functions. A plant homologue of RanBPM has not yet been characterized.

Plant Aurora kinases play a role in maintenance of primary meristems and control of endoreduplication.

• The conserved family of Aurora kinases has multiple functions during mitosis. The roles of plant Aurora kinases have been characterized using inhibitor treatments. • We down-regulated Aurora kinases in Arabidopsis thaliana using RNA interference (RNAi).

A nodulin/glutamine synthetase-like fusion protein is implicated in the regulation of root morphogenesis and in signalling triggered by flagellin.

The nodulin/glutamine synthetase-like protein (NodGS) that we identified proteomically in Arabidopsis thaliana is a fusion protein composed of an N-terminal amidohydrolase domain that shares homology with nodulins and a C-terminal domain of prokaryotic glutamine synthetase type I. The protein is homologous to the FluG protein, a morphogenetic factor in fungi. Although genes encoding NodGS homologues are present in many plant genomes, their products have not yet been characterized.

Polyamine metabolism during the cell cycle of synchronized tobacco BY-2 cell line.

The time courses of the contents of free, soluble and insoluble polyamine (PA) conjugates, PA biosynthetic and catabolic enzyme activities and mRNA levels of PA biosynthetic genes were monitored during the cell cycle of synchronized tobacco BY-2 cell line (Nicotiana tabacum L. cv. Bright Yellow 2).

Classical anticytokinins do not interact with cytokinin receptors but inhibit cyclin-dependent kinases.

Cytokinins are a class of plant hormones that regulate the cell cycle and diverse developmental and physiological processes. Several compounds have been identified that antagonize the effects of cytokinins. Based on structural similarities and competitive inhibition, it has been assumed that these anticytokinins act through a common cellular target, namely the cytokinin receptor.

Gamma-tubulin is essential for acentrosomal microtubule nucleation and coordination of late mitotic events in Arabidopsis.

Gamma-tubulin is required for microtubule (MT) nucleation at MT organizing centers such as centrosomes or spindle pole bodies, but little is known about its noncentrosomal functions. We conditionally downregulated gamma-tubulin by inducible expression of RNA interference (RNAi) constructs in Arabidopsis thaliana. Almost complete RNAi depletion of gamma-tubulin led to the absence of MTs and was lethal at the cotyledon stage.

Expression of a nondegradable cyclin B1 affects plant development and leads to endomitosis by inhibiting the formation of a phragmoplast.

In plants after the disassembly of mitotic spindle, a specific cytokinetic structure called the phragmoplast is built, and after cytokinesis, microtubules populate the cell cortex in an organized orientation that determines cell elongation and shape. Here, we show that impaired cyclin B1 degradation, resulting from a mutation within its destruction box, leads to an isodiametric shape of epidermal cells in leaves, stems, and roots and retarded growth of seedlings. Microtubules in these misshaped cells are grossly disorganized, focused around the nucleus, whereas they were entirely missing or abnormally organized along the cell cortex.

Plant gamma-tubulin interacts with alphabeta-tubulin dimers and forms membrane-associated complexes.

gamma-Tubulin is assumed to participate in microtubule nucleation in acentrosomal plant cells, but the underlying molecular mechanisms are still unknown. Here, we show that gamma-tubulin is present in protein complexes of various sizes and different subcellular locations in Arabidopsis and fava bean. Immunoprecipitation experiments revealed an association of gamma-tubulin with alphabeta-tubulin dimers.

A plant cyclin B2 is degraded early in mitosis and its ectopic expression shortens G2-phase and alleviates the DNA-damage checkpoint.

Mitotic progression is timely regulated by the accumulation and degradation of A- and B-type cyclins. In plants, there are three classes of A-, and two classes of B-type cyclins, but their specific roles are not known. We have generated transgenic tobacco plants in which the ectopic expression of a plant cyclin B2 gene is under the control of a tetracycline-inducible promoter.

A topoisomerase II-dependent checkpoint in G2-phase plant cells can be bypassed by ectopic expression of mitotic cyclin B2.

DNA topoisomerase II is required for mitotic chromosome condensation and segregation. Here we characterize the effects of inhibiting DNA topoisomerase II activity in plant cells using the non-DNA damaging topoisomerase II inhibitor ICRF-193. We report that ICRF-193 abrogated chromosome condensation in cultured alfalfa (Medicago sativa L.