Comprehensive Network Analysis of Anther-Expressed Genes in Rice by the Combination of 33 Laser Microdissection and 143 Spatiotemporal Microarrays

Co-expression networks systematically constructed from large-scale transcriptome data reflect the interactions and functions of genes with similar expression patterns and are a powerful tool for the comprehensive understanding of biological events and mining of novel genes. In Arabidopsis (a model dicot plant), high-resolution co-expression networks have been constructed from very large microarray datasets and these are publicly available as online information resources. However, the available transcriptome data of rice (a model monocot plant) have been limited so far, making it difficult for rice researchers to achieve reliable co-expression analysis. In this study, we performed co-expression network analysis by using combined 44 K agilent microarray datasets of rice, which consisted of 33 laser microdissection (LM)-microarray datasets of anthers, and 143 spatiotemporal transcriptome datasets deposited in RicexPro. The entire data of the rice co-expression network, which was generated from the 176 microarray datasets by the Pearson correlation coefficient (PCC) method with the mutual rank (MR)-based cut-off, contained 24,258 genes and 60,441 genes pairs. Using these datasets, we constructed high-resolution co-expression subnetworks of two specific biological events in the anther, “meiosis” and “pollen wall synthesis”. The meiosis network contained many known or putative meiotic genes, including genes related to meiosis initiation and recombination. In the pollen wall synthesis network, several candidate genes involved in the sporopollenin biosynthesis pathway were efficiently identified. Hence, these two subnetworks are important demonstrations of the efficiency of co-expression network analysis in rice. Our co-expression analysis included the separated transcriptomes of pollen and tapetum cells in the anther, which are able to provide precise information on transcriptional regulation during male gametophyte development in rice. The co-expression network data presented here is a useful resource for rice researchers to elucidate important and complex biological events

Topological phase-fluctuations, amplitude fluctuations, and criticality in extreme type-II superconductors

We study the effect of critical fluctuations on the $(B,T)$ phase diagram in extreme type-II superconductors in zero and finite magnetic field using large-scale Monte Carlo simulations on the Ginzburg-Landau model in a frozen gauge approximation. We show that a vortex-loop unbinding gives a correct picture of the zero field superconducting-normal transition even in the presence of amplitude fluctuations, which are far from being critical at $T_c$. We extract critical exponents of the dual model by studying the topological excitations of the original model. From the vortex-loop distribution function we extract the anomalous dimension of the dual field $\eta \simeq -0.18$, and conclude that the charged Ginzburg-Landau model and the neutral 3DXY model belong to different universality classes. We find are two distinct scaling regimes for the vortex-line lattice melting line: a high-field scaling regime and a distinct low-field 3DXY critical scaling regime. We also find indications of an abrupt change in the connectivity of the vortex-tangle in the vortex liquid along a line $T_L \geq T_M$. This is the finite field counter-part of the zero-field vortex-loop blowout. Which at low enough fields appears to coincide with $T_M$. Here, a description of the vortex system only in terms of field induced vortex lines is inadequate at and above the VLL melting temperature.Comment: 30 pages, 14 figure

Peak effect, vortex-lattice melting-line and order - disorder transition in conventional and high-T superconductors

We investigate the order - disorder transition line from a Bragg glass to an amorphous vortex glass in the H-T phase diagram of three-dimensional type-II superconductors with account of both pinning-caused and thermal fluctuations of the vortex lattice. Our approach is based on the Lindemann criterion and on results of the collective pinning theory and generalizes previous work of other authors. It is shown that the shapes of the order - disorder transition line and the vortex lattice melting curve are determined only by the Ginzburg number, which characterizes thermal fluctuations, and by a parameter which describes the strength of the quenched disorder in the flux-line lattice. In the framework of this unified approach we obtain the H-T phase diagrams for both conventional and high-Tc superconductors. Several well-known experimental results concerning the fishtail effect and the phase diagram of high-Tc superconductors are naturally explained by assuming that a peak effect in the critical current density versus H signalizes the order - disorder transition line in superconductors with point defects.Comment: 15 pages including 11 figure

A Novel RNA-Recognition-Motif Protein Is Required for Premeiotic G1/S-Phase Transition in Rice (Oryza sativa L.)

The molecular mechanism for meiotic entry remains largely elusive in flowering plants. Only Arabidopsis SWI1/DYAD and maize AM1, both of which are the coiled-coil protein, are known to be required for the initiation of plant meiosis. The mechanism underlying the synchrony of male meiosis, characteristic to flowering plants, has also been unclear in the plant kingdom. In other eukaryotes, RNA-recognition-motif (RRM) proteins are known to play essential roles in germ-cell development and meiosis progression. Rice MEL2 protein discovered in this study shows partial similarity with human proline-rich RRM protein, deleted in Azoospermia-Associated Protein1 (DAZAP1), though MEL2 also possesses ankyrin repeats and a RING finger motif. Expression analyses of several cell-cycle markers revealed that, in mel2 mutant anthers, most germ cells failed to enter premeiotic S-phase and meiosis, and a part escaped from the defect and underwent meiosis with a significant delay or continued mitotic cycles. Immunofluorescent detection revealed that T7 peptide-tagged MEL2 localized at cytoplasmic perinuclear region of germ cells during premeiotic interphase in transgenic rice plants. This study is the first report of the plant RRM protein, which is required for regulating the premeiotic G1/S-phase transition of male and female germ cells and also establishing synchrony of male meiosis. This study will contribute to elucidation of similarities and diversities in reproduction system between plants and other species

Bond-disordered spin systems: Theory and application to doped high-Tc compounds

We examine the stability of magnetic order in a classical Heisenberg model with quenched random exchange couplings. This system represents the spin degrees of freedom in high-$T_\textrm{c}$ compounds with immobile dopants. Starting from a replica representation of the nonlinear $\sigma$-model, we perform a renormalization-group analysis. The importance of cumulants of the disorder distribution to arbitrarily high orders necessitates a functional renormalization scheme. From the renormalization flow equations we determine the magnetic correlation length numerically as a function of the impurity concentration and of temperature. From our analysis follows that two-dimensional layers can be magnetically ordered for arbitrarily strong but sufficiently diluted defects. We further consider the dimensional crossover in a stack of weakly coupled layers. The resulting phase diagram is compared with experimental data for La$_{2-x}$Sr$_x$CuO$_4$.Comment: 12 pages, 5 figure

Hydrocarbon productivities in different Botryococcus strains: comparative methods in product quantification

Six different strains of the green microalgae Botryococcus belonging to the A-race or B-race, accumulating alkadiene or botryococcene hydrocarbons, respectively, were compared for biomass and hydrocarbon productivities. Biomass productivity was assessed gravimetrically upon strain growth in the laboratory under defined conditions. Hydrocarbon productivities were measured by three different and independent experimental approaches, including density equilibrium of the intact cells and micro-colonies, spectrophotometric analysis of hydrocarbon extracts, and gravimetric quantitation of eluted hydrocarbons. All three hydrocarbon-quantitation methods yielded similar results for each of the strains examined. The B-race microalgae Botryococcus braunii var. Showa and Kawaguchi-1 constitutively accumulated botryococcene hydrocarbons equivalent to 30% and 20%, respectively, of their overall biomass. The A-race microalgae Botryococcus braunii, varieties Yamanaka, UTEX 2441 and UTEX LB572 constitutively accumulated alkadiene hydrocarbons ranging from 14% to 13% and 10% of their overall biomass, respectively. Botryococcus sudeticus (UTEX 2629), a morphologically different green microalga, had the lowest hydrocarbon accumulation, equal to about 3% of its overall biomass. Results validate the density equilibrium and spectrophotometric analysis methods in the quantitation of botryococcene-type hydrocarbons. These analytical advances will serve in the screening and selection of B. braunii and of other microalgae in efforts to identify those having a high hydrocarbon content for use in commercial applications

Transcriptome Analysis of the Arabidopsis Megaspore Mother Cell Uncovers the Importance of RNA Helicases for Plant Germline Development

Germ line specification is a crucial step in the life cycle of all organisms. For sexual plant reproduction, the megaspore mother cell (MMC) is of crucial importance: it marks the first cell of the plant “germline” lineage that gets committed to undergo meiosis. One of the meiotic products, the functional megaspore, subsequently gives rise to the haploid, multicellular female gametophyte that harbours the female gametes. The MMC is formed by selection and differentiation of a single somatic, sub-epidermal cell in the ovule. The transcriptional network underlying MMC specification and differentiation is largely unknown. We provide the first transcriptome analysis of an MMC using the model plant Arabidopsis thaliana with a combination of laser-assisted microdissection and microarray hybridizations. Statistical analyses identified an over-representation of translational regulation control pathways and a significant enrichment of DEAD/DEAH-box helicases in the MMC transcriptome, paralleling important features of the animal germline. Analysis of two independent T-DNA insertion lines suggests an important role of an enriched helicase, MNEME (MEM), in MMC differentiation and the restriction of the germline fate to only one cell per ovule primordium. In heterozygous mem mutants, additional enlarged MMC-like cells, which sometimes initiate female gametophyte development, were observed at higher frequencies than in the wild type. This closely resembles the phenotype of mutants affected in the small RNA and DNA-methylation pathways important for epigenetic regulation. Importantly, the mem phenotype shows features of apospory, as female gametophytes initiate from two non-sister cells in these mutants. Moreover, in mem gametophytic nuclei, both higher order chromatin structure and the distribution of LIKE HETEROCHROMATIN PROTEIN1 were affected, indicating epigenetic perturbations. In summary, the MMC transcriptome sets the stage for future functional characterization as illustrated by the identification of MEM, a novel gene involved in the restriction of germline fate

Deficient activation of CD95 (APO-1/ Fas)-mediated apoptosis: a potential factor of multidrug resistance in human renal cell carcinoma

The pronounced resistance of human renal cell carcinoma (RCC) to anticancer-induced apoptosis has primarily been related to the expression of P-glycoprotein and effective drug detoxification mechanisms. Because the CD95 system has recently been identified as a key mediator of anticancer drug-induced apoptosis, we analysed the contribution of the CD95 system to chemotherapy-induced apoptosis in four newly established RCC cell lines. Here, we demonstrate that all RCC cell lines expressed CD95-receptor and -ligand. Exposure to agonistic anti-CD95 antibodies resulted in induction of apoptosis and significant (P< 0.05) reduction of cell number in three out of four cell lines, indicating that the essential components for CD95-mediated apoptosis were present and functionally intact in the majority of these RCC cell lines. Moreover, treatment of cultures with bleomycin or topotecan, a novel topoisomerase I inhibitor with little substrate affinity for P-glycoprotein, led to induction of apoptosis and significant (P< 0.05) dose-dependent reduction of cell number in all RCC cell lines. Both anticancer drugs also induced upregulation of CD95 ligand expression in all cell lines. Additionally, augmentation of CD95 receptor expression was found in three RCC cell lines, including one p53-mutated cell line, whereas another p53-mutated cell line showed no or only a weak CD95 receptor upregulation after exposure to topotecan or bleomycin, respectively. Despite this upregulation of CD95 receptor and ligand, antagonistic antibodies directed against CD95 receptors or ligands could not inhibit induction of apoptosis by topotecan and bleomycin in any cell line. Thus, although a functionally intact CD95 signalling cascade is present in most RCC cell lines, the anticancer drugs topotecan and bleomycin that induce upregulation of CD95 receptor and ligand fail to effectively activate CD95-mediated apoptosis. This deficient activation of CD95-mediated apoptosis might be an important additional factor for the multidrug resistance phenotype of human RCCs. © 2000 Cancer Research Campaig

The Role of MMP7 and Its Cross-Talk with the FAS/FASL System during the Acquisition of Chemoresistance to Oxaliplatin

Background: The efficacy of oxaliplatin in cancer chemotherapy is limited by the development of drug resistance. MMP7 has been related to the loss of tumor cell response to cytotoxic agents although the exact mechanism is not fully understood. Moreover, MMP7 is an independent prognosis factor for survival in patients with colorectal cancer. The aim of the present study was to analyze the role of MMP7 and its cross-talk with the Fas/FasL system during the acquisition of oxaliplatin resistance in colon cancer cells. Principal Findings: For this purpose we have developed three different oxaliplatin-resistant cell lines (RHT29, RHCT116 p53+/+, RHCT116 p53−/−) from the parental HT29, HCT116 p53+/+ and HCT116 p53−/− colon cancer cells. MMP7 basal expression was higher in the resistant compared to the parental cell lines. MMP7 was also upregulated by oxaliplatin in both HT29 (p53 mutant) and RHCT116 p53−/− but not in the RHCT116 p53+/+. Inhibition of MMP by 1,10-phenantroline monohydrate or siRNA of MMP7 restores cell sensitivity to oxaliplatin-induced apoptosis in both HT29 and RHCT116 p53−/− but not in the RHCT116 p53+/+. Some of these effects are caused by alterations in Fas receptor. Fas is upregulated by oxaliplatin in colon cancer cells, however the RHT29 cells treated with oxaliplatin showed a 3.8-fold lower Fas expression at the cell surface than the HT29 cells. Decrease of Fas at the plasma membrane seems to be caused by MMP7 since its inhibition restores Fas levels. Moreover, functional analysis of Fas demonstrates that this receptor was less potent in inducing apoptosis in RHT29 cells and that its activation induces MAPK signaling in resistant cells. Conclusions: Taking together, these results suggest that MMP7 is related to the acquisition of oxaliplatin-resistance and that its inhibition restores drug sensitivity by increasing Fas receptor. Furthermore, Fas undergoes a change in its functionality in oxaliplatin-resistant cells inducing survival pathways instead of apoptotic signals