The Gene Space database.

The hemiparasitic flowering plant (European mistletoe) is known for its very special life cycle, extraordinary biochemical properties, and extremely large genome. The size of its genome is estimated to be 30 times larger than the human genome and 600 times larger than the genome of the model plant . To achieve insights into the Gene Space of the genome, which is defined as the space including and surrounding protein-coding regions, a transcriptome project based on PacBio sequencing has recently been conducted.

The gene space of European mistletoe (Viscum album).

European mistletoe (Viscum album) is a hemiparasitic flowering plant that is known for its very special life cycle and extraordinary biochemical properties. Particularly, V. album has an unusual mode of cellular respiration that takes place in the absence of mitochondrial complex I.

Insights into the complex role of GRAS transcription factors in the arbuscular mycorrhiza symbiosis.

To improve access to limiting nutrients, the vast majority of land plants forms arbuscular mycorrhizal (AM) symbioses with Glomeromycota fungi. We show here that AM-related GRAS transcription factors from different subgroups are upregulated during a time course of mycorrhization. Based on expression studies in mutants defective in arbuscule branching (ram1-1, with a deleted MtRam1 GRAS transcription factor gene) or in the formation of functional arbuscules (pt4-2, mutated in the phosphate transporter gene MtPt4), we demonstrate that the five AM-related GRAS transcription factor genes MtGras1, MtGras4, MtGras6, MtGras7, and MtRad1 can be differentiated by their dependency on MtRAM1 and MtPT4, indicating that the network of AM-related GRAS transcription factors consists of at least two regulatory modules.

The mycorrhiza-dependent defensin MtDefMd1 of Medicago truncatula acts during the late restructuring stages of arbuscule-containing cells.

Different symbiotic and pathogenic plant-microbe interactions involve the production of cysteine-rich antimicrobial defensins. In Medicago truncatula, the expression of four MtDefMd genes, encoding arbuscular mycorrhiza-dependent defensins containing an N-terminal signal peptide and exhibiting some differences to non-symbiotic defensins, raised over the time of fungal colonization. Whereas the MtDefMd1 and MtDefMd2 promoters were inactive in cells containing young arbuscules, cells with fully developed arbuscules displayed different levels of promoter activities, indicating an up-regulation towards later stages of arbuscule formation.

Pre-announcement of symbiotic guests: transcriptional reprogramming by mycorrhizal lipochitooligosaccharides shows a strict co-dependency on the GRAS transcription factors NSP1 and RAM1.

Background: More than 80 % of all terrestrial plant species establish an arbuscular mycorrhiza (AM) symbiosis with Glomeromycota fungi. This plant-microbe interaction primarily improves phosphate uptake, but also supports nitrogen, mineral, and water aquisition. During the pre-contact stage, the AM symbiosis is controled by an exchange of diffusible factors from either partner.

Transcriptional responses toward diffusible signals from symbiotic microbes reveal MtNFP- and MtDMI3-dependent reprogramming of host gene expression by arbuscular mycorrhizal fungal lipochitooligosaccharides.

The formation of root nodules and arbuscular mycorrhizal (AM) roots is controlled by a common signaling pathway including the calcium/calmodulin-dependent kinase Doesn't Make Infection3 (DMI3). While nodule initiation by lipochitooligosaccharide (LCO) Nod factors is well characterized, diffusible AM fungal signals were only recently identified as sulfated and nonsulfated LCOs. Irrespective of different outcomes, the perception of symbiotic LCOs in Medicago truncatula is mediated by the LysM receptor kinase M.

Laser microdissection unravels cell-type-specific transcription in arbuscular mycorrhizal roots, including CAAT-box transcription factor gene expression correlating with fungal contact and spread.

Arbuscular mycorrhizae (AM) are the most widespread symbioses on Earth, promoting nutrient supply of most terrestrial plant species. To unravel gene expression in defined stages of Medicago truncatula root colonization by AM fungi, we here combined genome-wide transcriptome profiling based on whole mycorrhizal roots with real-time reverse transcription-PCR experiments that relied on characteristic cell types obtained via laser microdissection. Our genome-wide approach delivered a core set of 512 genes significantly activated by the two mycorrhizal fungi Glomus intraradices and Glomus mossae.

Purine receptors and Ca(2+) signalling in the human blood-brain barrier endothelial cell line hCMEC/D3.

The expression and physiology of purine receptors of the human blood-brain barrier endothelial cells were characterised by application of molecular biological, gene-silencing and Ca(2+)-imaging techniques to hCMEC/D3 cells. Reverse transcription polymerase chain reaction showed the expression of the G-protein-coupled receptors P2Y(2)-, P2Y(6)-, P2Y(11)- as well as the ionotropic P2X(4)-, P2X(5)- and P2X(7)-receptors. Fura-2 ratiometry revealed that adenosine triphosphate (ATP) or uridine triphosphate (UTP) mediated a change in the intracellular Ca(2+) concentration ([Ca(2+)](i)) from 150 to 300 nM in single cells.

Knockdown of the symbiotic sucrose synthase MtSucS1 affects arbuscule maturation and maintenance in mycorrhizal roots of Medicago truncatula.

The relevance of the symbiosis-induced Medicago truncatula sucrose synthase gene MtSucS1 for an efficient arbuscular mycorrhiza (AM) was studied using two independent antisense lines that displayed up to 10-fold reduced SucS1 levels in roots. Mycorrhizal MtSucS1-reduced lines exhibited an overall stunted aboveground growth under inorganic phosphorus limitation. Apart from a reduced plant height, shoot weight, and leaf development, a delayed flowering, resulting in a lower seed yield, was observed.

Differential gel electrophoresis (DIGE) to quantitatively monitor early symbiosis- and pathogenesis-induced changes of the Medicago truncatula root proteome.

Symbiosis- and pathogenesis-related early protein induction patterns in the model legume Medicago truncatula were analysed with two-dimensional differential gel electrophoresis. Two symbiotic soil microorganisms (Glomus intraradices, Sinorhizobium meliloti) were used in single infections and in combination with a secondary pathogenic infection by the oomycete Aphanomyces euteiches. Proteomic analyses performed 6 and 24h after inoculations led to identification of 87 differentially induced proteins which likely represent the M.

The signal peptide of the Medicago truncatula modular nodulin MtNOD25 operates as an address label for the specific targeting of proteins to nitrogen-fixing symbiosomes.

The nodule-specific MtNOD25 gene of the model legume Medicago truncatula encodes a modular nodulin composed of different repetitive modules flanked by distinct N- and C-termini. Although similarities are low with respect to all repetitive modules, both the N-terminal signal peptide (SP) and the C-terminus are highly conserved in modular nodulins from different legumes. On the cellular level, MtNOD25 is only transcribed in the infected cells of root nodules, and this activation is mediated by a 299-bp minimal promoter containing an organ-specific element.

Evidence for the involvement in nodulation of the two small putative regulatory peptide-encoding genes MtRALFL1 and MtDVL1.

Nod factors are key bacterial signaling molecules regulating the symbiotic interaction between bacteria known as rhizobia and leguminous plants. Studying plant host genes whose expression is affected by Nod factors has given insights into early symbiotic signaling and development. Here, we used a double supernodulating mutant line that shows increased sensitivity to Nod factors to study the Nod factor-regulated transcriptome.

Evidence for transcriptional and post-translational regulation of sucrose synthase in pea nodules by the cellular redox state.

Nitrogen fixation (NF) in legume nodules is very sensitive to environmental constraints. Nodule sucrose synthase (SS; EC 2.4.

Antisense repression of the Medicago truncatula nodule-enhanced sucrose synthase leads to a handicapped nitrogen fixation mirrored by specific alterations in the symbiotic transcriptome and metabolome.

We analyzed the role of the sucrose (Suc) synthase MtSucS1 during nodulation of the model legume Medicago truncatula, integrating data for the developmental, transcriptional, and metabolic processes affected downstream of an impaired Suc cleavage in root nodules. To reduce carbohydrate supply to nodule tissues, transgenic plants expressing a p35S-driven MtSucS1-antisense fusion were constructed. These plants displayed an up to 90% reduction of MtSucS1 proteins in roots and nodules.

Development of bioinformatic tools to support EST-sequencing, in silico- and microarray-based transcriptome profiling in mycorrhizal symbioses.

The great majority of terrestrial plants enters a beneficial arbuscular mycorrhiza (AM) or ectomycorrhiza (ECM) symbiosis with soil fungi. In the SPP 1084 "MolMyk: Molecular Basics of Mycorrhizal Symbioses", high-throughput EST-sequencing was performed to obtain snapshots of the plant and fungal transcriptome in mycorrhizal roots and in extraradical hyphae. To focus activities, the interactions between Medicago truncatula and Glomus intraradices as well as Populus tremula and Amanita muscaria were selected as models for AM and ECM symbioses, respectively.

Identification and expression regulation of symbiotically activated legume genes.

Legume plants are able to enter two different endosymbioses with soil prokaryotes and soil fungi, leading to nitrogen-fixing root nodules and to arbuscular mycorrhiza (AM), respectively. We applied in silico and microarray-based transcriptome profiling approaches to uncover the transcriptome of developing root nodules and AM roots of the model legume Medicago truncatula. Several hundred genes were found to be activated in different stages of either symbiosis, with almost 100 genes being co-induced during nodulation and in arbuscular mycorrhiza.

Transcriptional snapshots provide insights into the molecular basis of arbuscular mycorrhiza in the model legume Medicago truncatula.

The arbuscular mycorrhizal (AM) association between terrestrial plants and soil fungi of the phylum Glomeromycota is the most widespread beneficial plant-microbe interaction on earth. In the course of the symbiosis, fungal hyphae colonise plant roots and supply limiting nutrients, in particular phosphorus, in exchange for carbon compounds. Owing to the obligate biotrophy of mycorrhizal fungi and the lack of genetic systems to study them, targeted molecular studies on AM symbioses proved to be difficult.

Overlaps in the transcriptional profiles of Medicago truncatula roots inoculated with two different Glomus fungi provide insights into the genetic program activated during arbuscular mycorrhiza.

Arbuscular mycorrhiza (AM) is a widespread symbiotic association between plants and fungal microsymbionts that supports plant development under nutrient-limiting and various stress conditions. In this study, we focused on the overlapping genetic program activated by two commonly studied microsymbionts in addition to identifying AM-related genes. We thus applied 16,086 probe microarrays to profile the transcriptome of the model legume Medicago truncatula during interactions with Glomus mosseae and Glomus intraradices and specified a total of 201 plant genes as significantly coinduced at least 2-fold, with more than 160 being reported as AM induced for the first time.

The promoter of the leghaemoglobin gene VfLb29: functional analysis and identification of modules necessary for its activation in the infected cells of root nodules and in the arbuscule-containing cells of mycorrhizal roots.

In this study the further characterization of the Vicia faba leghaemoglobin promoter pVfLb29 is presented that was previously shown to be specifically active in the infected cells of root nodules and in arbuscule-containing cells of mycorrhizal roots. Using promoter studies in transgenic hairy roots of the Pisum sativum mutant RisNod24, disabled in the formation of functional arbuscules, VfLb29 promoter activity is assigned to later stages of arbuscule development. In order to narrow down the regions containing cis-acting elements of pVfLb29, the activity of five VfLb29 promoter deletions (-797/-31 to -175/-31 in relation to the start codon) fused to the gusAint coding region were tested in transgenic V.

Two genes encoding different truncated hemoglobins are regulated during root nodule and arbuscular mycorrhiza symbioses of Medicago truncatula.

The MtTrHb1 and MtTrHb2 genes of the model legume Medicago truncatula Gaertn. encode proteins homologous to truncated hemoglobins (TrHb) from plants and a range of different microorganisms. Induction of MtTrHb1 in root nodules and expression of MtTrHb2 in root nodules, as well as in mycorrhizal roots, were shown by quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR).

Transcriptome profiling in root nodules and arbuscular mycorrhiza identifies a collection of novel genes induced during Medicago truncatula root endosymbioses.

Transcriptome profiling based on cDNA array hybridizations and in silico screening was used to identify Medicago truncatula genes induced in both root nodules and arbuscular mycorrhiza (AM). By array hybridizations, we detected several hundred genes that were upregulated in the root nodule and the AM symbiosis, respectively, with a total of 75 genes being induced during both interactions. The second approach based on in silico data mining yielded several hundred additional candidate genes with a predicted symbiosis-enhanced expression.

Expression profiling in Medicago truncatula identifies more than 750 genes differentially expressed during nodulation, including many potential regulators of the symbiotic program.

In this study, we describe a large-scale expression-profiling approach to identify genes differentially regulated during the symbiotic interaction between the model legume Medicago truncatula and the nitrogen-fixing bacterium Sinorhizobium meliloti. Macro- and microarrays containing about 6,000 probes were generated on the basis of three cDNA libraries dedicated to the study of root symbiotic interactions. The experiments performed on wild-type and symbiotic mutant material led us to identify a set of 756 genes either up- or down-regulated at different stages of the nodulation process.

Construction and validation of cDNA-based Mt6k-RIT macro- and microarrays to explore root endosymbioses in the model legume Medicago truncatula.

To construct macro- and microarray tools suitable for expression profiling in root endosymbioses of the model legume Medicago truncatula, we PCR-amplified a total of 6048 cDNA probes representing genes expressed in uninfected roots, mycorrhizal roots and young root nodules [Nucleic Acids Res. 30 (2002) 5579]. Including additional probes for either tissue-specific or constitutively expressed control genes, 5651 successfully amplified gene-specific probes were used to grid macro- and to spot microarrays designated Mt6k-RIT (M.

The Medicago truncatula sucrose synthase gene MtSucS1 is activated both in the infected region of root nodules and in the cortex of roots colonized by arbuscular mycorrhizal fungi.

The MtSucS1 gene encodes a sucrose synthase (EC 2.4.1.

The promoter of the Vicia faba L. gene VfEnod12 encoding an early nodulin is active in cortical cells and nodule primordia of transgenic hairy roots of Vicia hirsuta as well as in the prefixing zone II of mature transgenic V. hirsuta root nodules.

A full-length cDNA encoding the Vicia faba L. early nodulin VfEnod12 was isolated. The deduced protein sequence specified a 90 amino acid protein with a MW of 10206 and contained a putative signal peptide sequence followed by PPX(3) repeats characteristic of Enod12 proteins.