SPX1 is a phosphate-dependent inhibitor of Phosphate Starvation Response 1 in Arabidopsis.

To cope with growth in low-phosphate (Pi) soils, plants have evolved adaptive responses that involve both developmental and metabolic changes. Phosphate Starvation Response 1 (PHR1) and related transcription factors play a central role in the control of Pi starvation responses (PSRs). How Pi levels control PHR1 activity, and thus PSRs, remains to be elucidated. Here, we identify a direct Pi-dependent inhibitor of PHR1 in Arabidopsis, SPX1, a nuclear protein that shares the SPX domain with yeast Pi sensors and with several Pi starvation signaling proteins from plants. Double mutation of SPX1 and of a related gene, SPX2, resulted in molecular and physiological changes indicative of increased PHR1 activity in plants grown in Pi-sufficient conditions or after Pi refeeding of Pi-starved plants but had only a limited effect on PHR1 activity in Pi-starved plants. These data indicate that SPX1 and SPX2 have a cellular Pi-dependent inhibitory effect on PHR1. Coimmunoprecipitation assays showed that the SPX1/PHR1 interaction in planta is highly Pi-dependent. DNA-binding and pull-down assays with bacterially expressed, affinity-purified tagged SPX1 and ΔPHR1 proteins showed that SPX1 is a competitive inhibitor of PHR1 binding to its recognition sequence, and that its efficiency is highly dependent on the presence of Pi or phosphite, a nonmetabolizable Pi analog that can repress PSRs. The relative strength of the SPX1/PHR1 interaction is thus directly influenced by Pi, providing a link between Pi perception and signaling.

Molecular genetic basis of pod corn (Tunicate maize).

Pod corn is a classic morphological mutant of maize in which the mature kernels of the cob are covered by glumes, in contrast to generally grown maize varieties in which kernels are naked. Pod corn, known since pre-Columbian times, is the result of a dominant gain-of-function mutation at the Tunicate (Tu) locus. Some classic articles of 20th century maize genetics reported that the mutant Tu locus is complex, but molecular details remained elusive. Here, we show that pod corn is caused by a cis-regulatory mutation and duplication of the ZMM19 MADS-box gene. Although the WT locus contains a single-copy gene that is expressed in vegetative organs only, mutation and duplication of ZMM19 in Tu lead to ectopic expression of the gene in the inflorescences, thus conferring vegetative traits to reproductive organs.

Whole Body Vibration Exercise Protocol versus a Standard Exercise Protocol after ACL Reconstruction: A Clinical Randomized Controlled Trial with Short Term Follow-Up.

The suitability and effectiveness of whole body vibration (WBV) exercise in rehabilitation after injury of the anterior cruciate ligament (ACL) was studied using a specially designed WBV protocol. We wanted to test the hypothesis if WBV leads to superior short term results regarding neuromuscular performance (strength and coordination) and would be less time consuming than a current standard muscle strengthening protocol. In this prospective randomized controlled clinical trial, forty patients who tore their ACL and underwent subsequent ligament reconstruction were enrolled. Patients were randomized to the whole body vibration (n=20) or standard rehabilitation exercise protocol (n=20). Both protocols started in the 2(nd) week after surgery. Isometric and isokinetic strength measurements, clinical assessment, Lysholm score, neuromuscular performance were conducted weeks 2, 5, 8 and 11 after surgery. Time spent for rehabilitation exercise was reduced to less than a half in the WBV group. There were no statistically significant differences in terms of clinical assessment, Lysholm score, isokinetic and isometric strength. The WBV group displayed significant better results in the stability test. In conclusion, preliminary data indicate that our whole body vibration muscle exercise protocol seems to be a good alternative to a standard exercise program in ACL-rehabilitation. Despite of its significant reduced time requirement it is at least equally effective compared to a standard rehabilitation protocol. Key pointsIn this prospective randomized controlled clinical trial, we tested the hypothesis if WBV leads to superior short term results regarding neuromuscular performance (strength and coordination) and would be less time consuming than a current standard muscle strengthening protocol in forty patients who underwent ACL reconstruction.Time spent for rehabilitation exercise was reduced to less than a half in the WBV group as compared to the standard exercise group. Both protocols showed no differences regarding clinical assessment, Lysholm score, isokinetic and isometric strength.Despite a more than 50% reduction in time spent for exercise sessions, the WBV group achieved significant better results in the stability test.In conclusion, the presented WBV program can be considered as a practical alternative to a standard exercise program during ACL-rehabilitation.

Microcirculation Under an Elastic Bandage During Rest and Exercise - Preliminary Experience With the Laser-Doppler Spectrophotometry System O2C.

There is an abundace of studies on the influence of rest and exercise as well as external compression on cutaneous, subcutaneous and muscle tissue blood flow using different measurement techniques. As a novel approach, we simultaneously examined the influence of a custom- made elastic thigh bandage on cutaneous and subcutaneous venous blood oxygenation (SO2), postcapillary venous filling pressures (rHb) and blood flow (flow) using the non-invasive laser- Doppler spectrophotometry system "Oxygen-to-see(O2C)". Parameters were obtained in 20 healthy volunteers in 2 mm and 8 mm tissue depth during rest, 5 and 10 minutes of moderate bicycle exercise following a 10-minute recovery period. Without the bandage, results matched the known physiological changes indicating higher blood backflow from superficial and deep veins. Underneath the elastic bandage, we observed lower post-capillary filling pressures during exercise. However, after the bandage was removed in the post-exercise period, all obtained parameters of microcirculation remained increased, indicating a higher amount of local venous blood volume in this area. Our observations might be the result of external compression, thermoregulatory and exercise-dependent vascular mechanisms. With the O2C device, a promising new non- invasive technique of measuring local microcirculation in soft tissue exists. This study gives new insights in the field of non-invasive diagnostics with special regard to the influence of elastic bandages on local microcirculation. Key PointsIt can be demonstrated that a novel non-invasive laser-Doppler spectrophotometry system allows the determination of capillary-venous microcirculation in an in-vivo study during exercise-rest cycles.The results received with this technique indicate that a) without an elastic thigh bandage, turnover rates of capillary and post-capillary microperfusion in skin and subcutaneous fat tissue increase under physical exertion, b) skin blood flow decreases while subcutaneous blood flow remained constant in the subsequent recovery phase. While wearing the bandage, c) venous back flow during exercise is increased, whereas d) in the recovery phase, microcirculation remained increased in both tissue depths after removing the bandage.In conclusion, the elastic bandage has a negative impact on local microcirculation and capillary-venous back flow, which is possibly due to a displacement of blood volume into the deep venous system and heat accumulation impairing the thermoregulatory response at the same time.

ERAMOSA controls lateral branching in snapdragon.

Plant forms display a wide variety of architectures, depending on the number of lateral branches, internode elongation and phyllotaxy. These are in turn determined by the number, the position and the fate of the Axillary Meristems (AMs). Mutants that affect AM determination during the vegetative phase have been isolated in several model plants. Among these genes, the GRAS transcription factor LATERAL SUPPRESSOR (Ls) plays a pivotal role in AM determination during the vegetative phase. Hereby we characterize the phylogenetic orthologue of Ls in Antirrhinum, ERAMOSA (ERA). Our data supported ERA control of AM formation during both the vegetative and the reproductive phase in snapdragon. A phylogenetic analysis combined with an analysis of the synteny of Ls in several species strongly supported the hypothesis that ERA is a phylogenetic orthologue of Ls, although it plays a broader role. During the reproductive phase ERA promotes the establishment of the stem niche at the bract axis but, after the reproductive transition, it is antagonized by the MADS box transcription factor SQUAMOSA (SQUA). Surprisingly double mutant era squa plants display a squa phenotype developing axillary meristems, which can eventually turn into inflorescences or flowers.

The DefH9-iaaM-containing construct efficiently induces parthenocarpy in cucumber.

Parthenocarpy (seedless fruits) is a desirable trait that has been achieved in many plant cultivars. We generated parthenocarpic cucumber fruits by introducing the chimeric DefH9-iaaM construct into the cucumber genome using an Agrobacterium tumefaciens-mediated protocol. The construct consists of the DefH9 promoter from Antirrhinum majus and the iaaM coding sequence from Pseudomonas syringae. Transgenic plants were obtained from nine independent transformation events: half of these were tetraploid and did not produce seeds following self-pollination, while the remaining half were capable of displaying parthenocarpy in the subsequent reproductive generation. Of the fruits produced by the transgenic lines, 70-90% were parthenocarpic. The segregation of the marker gene in the transgenic T(1) progeny indicated single gene inheritance. The seed set in the transgenic lines and their F(1) hybrids was lower than in the non-transgenic control plants. Some of the methodological details and the practical significance of the results are discussed.

PFMAGO, a MAGO NASHI-like factor, interacts with the MADS-domain protein MPF2 from Physalis floridana.

MADS-domain proteins serve as regulators of plant development and often form dimers and higher order complexes to function. Heterotopic expression of MPF2, a MADS-box gene, in reproductive tissues is a key component in the evolution of the inflated calyx syndrome in Physalis, but RNAi studies demonstrate that MPF2 has also acquired a role in male fertility in Physalis floridana. Using the yeast 2-hybrid system, we have now identified numerous MPF2-interacting MADS-domain proteins from Physalis, including homologs of SOC1, AP1, SEP1, SEP3, AG, and AGL6. Among the many non-MADS-domain proteins recovered was a homolog of MAGO NASHI, a highly conserved RNA-binding protein known to be involved in many developmental processes including germ cell differentiation. Two MAGO genes, termed P. floridana mago nashi1 (PFMAGO1) and PFMAGO2, were isolated from P. floridana. Both copies were found to be coexpressed in leaves, fruits, and, albeit at lower level, also in roots, stems, and flowers. DNA sequence analysis revealed that, although the coding sequences of the 2 genes are highly conserved, they differ substantially in their intron and promoter sequences. Two-hybrid screening of a Physalis expression library with both PFMAGO1 and PFMAGO2 as baits yielded numerous gene products, including an Y14-like protein. Y14 is an RNA-binding protein that forms part of various "gene expression machines." The function of MPF2 and 2 PFMAGO proteins in ensuring male fertility and evolution of calyx development in Physalis is discussed.

ROSINA (RSI) is part of a CACTA transposable element, TamRSI, and links flower development to transposon activity.

ROSINA (RSI) was isolated as a DNA binding factor able to bind to the CArG-box present in the promoter of the MADS-box gene DEFICIENS of Antirrhinum majus. The mosaic nature of RSI and its multi-copy presence in the A. majus genome indicated that RSI could be a part of a mobile genetic element. Here we show that RSI is a part of a CACTA transposable element system of A. majus, named TamRSI, which has evolved and is still evolving within the terminal inverted repeats (TIRs) of this CACTA transposon. Interestingly, RSI is always found in opposite orientation with respect to the transcription of a second gene present within the CACTA transposon, which encodes a putative TRANSPOSASE (TNP). This structural configuration has not yet been described for any member of the CACTA transposons superfamily. Internal deletion derivatives of the TamRSI produce aberrant RSI transcripts (RSI-ATs) that carry parts of the RSI RNA fused to parts of the TNP RNA. In addition, an intriguing seed phenotype shown by RNAi transgenic lines generated to silence RSI, relate TamRSI to epigenetic mechanisms and associate the control of flower development to transposon activity.

ROSINA (RSI), a novel protein with DNA-binding capacity, acts during floral organ development in Antirrhinum majus.

Petal and stamen identity of the Antirrhinum majus flower is under the genetic control of the floral homeotic gene DEFICIENS (DEF). To isolate factors involved in the regulation of DEF gene activity, a promoter segment of this B-function gene, containing cis-acting regulatory elements, was used to identify the novel trans-acting factor ROSINA (RSI). RSI does not show an extended similarity with any gene product present in the database. Rather RSI constitutes a protein that contains domains similar to known proteins from organisms of different phyla. The capacity of RSI to bind a sequence element of the DEF promoter, its spatial and temporal expression pattern together with the phenotype of RSI-RNAi interference plants as well as RSI over-expression in Arabidopsis thaliana suggest that RSI is a putative regulator of DEF gene activity in A. majus.

Identification and functional characterisation of ctr1, a Pleurotus ostreatus gene coding for a copper transporter.

Copper homeostasis is crucial for the maintenance of life. In lignin-degrading fungi, copper is essential for the phenol oxidase enzymes that provide this activity. In this paper we report the characterization of a gene (ctr1) coding for a copper transporter in the white rot fungus Pleurotus ostreatus. The gene was identified in a cDNA library constructed from 4-day-old vegetative mycelium grown in liquid culture. The results presented here demonstrate that: (1) ctr1 functionally complements the respiratory deficiency of a yeast mutant defective in copper transport, supporting the idea that the Ctr1 protein is itself a copper transporter; (2) transcription of ctr1 is detectable in P. ostreatus at all developmental stages and in all tissues (with the exception of lamellae), and is negatively regulated by the presence of copper in the culture medium; (3) ctr1 is a single-copy gene that maps to P. ostreatus linkage group III; and (4) the regulatory sequence elements found in the promoter of ctr1 are similar to those found in other copper-related genes described in other systems. These results provide the first description of a copper transporter in this white rot fungus and should be useful for further studies on copper metabolism in higher basidiomycetes.

INCOMPOSITA: a MADS-box gene controlling prophyll development and floral meristem identity in Antirrhinum.

INCOMPOSITA (INCO) is a MADS-box transcription factor and member of the functionally diverse StMADS11 clade of the MADS-box family. The most conspicuous feature of inco mutant flowers are prophylls initiated prior to first whorl sepals at lateral positions of the flower primordium. The developing prophylls physically interfere with subsequent floral organ development that results in aberrant floral architecture. INCO, which is controlled by SQUAMOSA, prevents prophyll formation in the wild type, a role that is novel among MADS-box proteins, and we discuss evolutionary implications of this function. Overexpression of INCO or SVP, a structurally related Arabidopsis MADS-box gene involved in the negative control of Arabidopsis flowering time, conditions delayed flowering in transgenic plants, suggesting that SVP and INCO have functions in common. Enhanced flowering of squamosa mutants in the inco mutant background corroborates this potential role of INCO as a floral repressor in Antirrhinum. One further, hitherto hidden, role of INCO is the positive control of Antirrhinum floral meristem identity. This is revealed by genetic interactions between inco and mutants of FLORICAULA, a gene that controls the inflorescence to floral transition, together with SQUAMOSA. The complex regulatory and combinatorial relations between INCO, FLORICAULA and SQUAMOSA are summarised in a model that integrates observations from molecular studies as well as analyses of expression patterns and genetic interactions.

Characterization of antirrhinum petal development and identification of target genes of the class B MADS box gene DEFICIENS.

The class B MADS box transcription factors DEFICIENS (DEF) and GLOBOSA (GLO) of Antirrhinum majus together control the organogenesis of petals and stamens. Toward an understanding of how the downstream molecular mechanisms controlled by DEF contribute to petal organogenesis, we conducted expression profiling experiments using macroarrays comprising >11,600 annotated Antirrhinum unigenes. First, four late petal developmental stages were compared with sepals. More than 500 ESTs were identified that comprise a large number of stage-specifically regulated genes and reveal a highly dynamic transcriptional regulation. For identification of DEF target genes that might be directly controlled by DEF, we took advantage of the temperature-sensitive def-101 mutant. To enhance the sensitivity of the profiling experiments, one petal developmental stage was selected, characterized by increased transcriptome changes that reflect the onset of cell elongation processes replacing cell division processes. Upon reduction of the DEF function, 49 upregulated and 52 downregulated petal target genes were recovered. Eight target genes were further characterized in detail by RT-PCR and in situ studies. Expression of genes responding rapidly toward an altered DEF activity is confined to different petal tissues, demonstrating the complexity of the DEF function regulating diverse basic processes throughout petal morphogenesis.