bluuebunny/crossref_metadata_2025_split huggingface dataset:
Acceleration of Aux/IAA proteolysis is specific for auxin and independent of AXR1
Authors: Zenser Nathan , Dreher Kate A. , Edwards Stephanie R. , Callis Judy 
Published: July 2003

SummaryAux/IAA proteins are short‐lived transcriptional regulators involved in auxin signaling. Using Aux/IAA luciferase (LUC) fusion proteins expressed in Arabidopsis thaliana, we previously showed that rapid degradation of these proteins requires conserved Aux/IAA domain II and that exogenous auxin accelerates their degradation. To further examine auxin‐mediated increases in proteolysis, the degradation of two other LUC fusion proteins, a non‐cleavable ubiquitin LUC fusion (UB1–72::LUC) and SAUR15::LUC was determined in vivo in seedlings. Their half‐lives were 20 ± 4 and 104 ± 10 min, respectively. SAUR15::LUC half‐life was not affected by pre‐incubation with 2,4‐D. Auxin did not have an equivalent effect on UB(1–72)::LUC steady‐state levels as compared to PsIAA6:LUC. LUC fused to an Aux/IAA domain II degraded more rapidly following auxin application, demonstrating that this region is sufficient for auxin‐mediated acceleration of proteolysis. Hormonal cross‐talk at the level of Aux/IAA proteolysis was examined. 1‐aminocyclopropane‐1‐carboxylic acid (ACC), benzyladenine (BA), abscisic acid (ABA), and brassinolide (BL) did not affect the degradation rate of IAA1::LUC, and gibberellic acid (GA3) and salicylic acid (SA) did not specifically affect the steady‐state levels of Aux/IAA::LUC proteins. An Aux/IAA::LUC transgene was crossed into the auxin resistant‐1 (axr1–12) background. In axr1–12, the half‐life of PsIAA6(1–73)::LUC increased 4.5‐fold, but proteolysis still accelerated in response to exogenous auxin. These data suggest that auxin is the only phytohormone that accelerates Aux/IAA proteolysis, and that this acceleration is specific for Aux/IAA proteins. In addition, AXR1 plays an important role in rapid basal proteolysis of Aux/IAA proteins, but is not required for auxin‐mediated acceleration of their degradation.
The soybean NRAMP homologue, GmDMT1, is a symbiotic divalent metal transporter capable of ferrous iron transport
Authors: Kaiser Brent N. , Moreau Sophie , Castelli Joanne , Thomson Rowena , Lambert Annie , Bogliolo Stéphanie , Puppo Alain , Day David A. 
Published: July 2003

SummaryIron is an important nutrient in N2‐fixing legume root nodules. Iron supplied to the nodule is used by the plant for the synthesis of leghemoglobin, while in the bacteroid fraction, it is used as an essential cofactor for the bacterial N2‐fixing enzyme, nitrogenase, and iron‐containing proteins of the electron transport chain. The supply of iron to the bacteroids requires initial transport across the plant‐derived peribacteroid membrane, which physically separates bacteroids from the infected plant cell cytosol. In this study, we have identified Glycine max divalent metal transporter 1 (GmDmt1), a soybean homologue of the NRAMP/Dmt1 family of divalent metal ion transporters. GmDmt1 shows enhanced expression in soybean root nodules and is most highly expressed at the onset of nitrogen fixation in developing nodules. Antibodies raised against a partial fragment of GmDmt1 confirmed its presence on the peribacteroid membrane (PBM) of soybean root nodules. GmDmt1 was able to both rescue growth and enhance 55Fe(II) uptake in the ferrous iron transport deficient yeast strain (fet3fet4). The results indicate that GmDmt1 is a nodule‐enhanced transporter capable of ferrous iron transport across the PBM of soybean root nodules. Its role in nodule iron homeostasis to support bacterial nitrogen fixation is discussed.
Rapid migration in gel filtration of the Cf‐4 and Cf‐9 resistance proteins is an intrinsic property of Cf proteins and not because of their association with high‐molecular‐weight proteins
Authors: Van der Hoorn Renier A. L. , Rivas Susana , Wulff Brande B. H. , Jones Jonathan D. G. , Joosten Matthieu H. A. J. 
Published: July 2003

SummaryGel filtration is frequently used to study the behaviour and composition of protein complexes. In previous studies, gel filtration analysis of solubilised membranes containing the tomato Cf‐4 and Cf‐9 resistance proteins indicated that these Cf proteins are present in an approximately 400‐ and 420‐kDa protein complex, respectively, which contains only one Cf molecule per complex, does not contain Rho‐related proteins, and does not alter in size upon elicitation. Here, we show that inactive Cf‐4 and Cf‐9 mutant proteins have a similar large apparent size upon gel filtration analysis. The size remains unaltered after pre‐treating the samples under harsh conditions, such as boiling with SDS and incubation in 6 m urea. A similar large apparent size was found for Cf‐4 and Cf‐9 isolated from SDS gel and for Cf‐9 expressed by insect cells. Therefore, the large apparent size observed in our studies appears to be an intrinsic property of the Cf proteins, rather than being caused by association with high‐molecular‐weight protein(s). Taken together, these results suggest that caution should be taken when interpreting data obtained from gel filtration of LRR‐containing proteins.
The Arabidopsis mutant dct is deficient in the plastidic glutamate/malate translocator DiT2
Authors: Renné Petra , Dreßen Uta , Hebbeker Ulrike , Hille Diana , Flügge Ulf‐Ingo , Westhoff Peter , Weber Andreas P. M. 
Published: July 2003

SummaryThe Arabidopsis mutant dicarboxylate transport (dct) is one of the classic mutants in the photorespiratory pathway. It requires high CO2 levels for survival. Physiologic and biochemical characterization of dct indicated that dct is deficient in the transport of dicarboxylates across the chloroplast envelope membrane. Hence, re‐assimilation of ammonia generated by the photorespiratory cycle is blocked. However, the defective gene in dct has not been identified at the molecular level. Here, we report on the molecular characterization of the defective gene in dct, on the complementation of the mutant phenotype with a wild‐type cDNA, and on the functional characterization of the gene product, DiT2, in a recombinant reconstituted system. Furthermore, we provide the kinetic constants of recombinant DiT1 and DiT2, and we discuss these data with respect to their functions in ammonia assimilation. Moreover, an analysis of the transcript levels of DiT1 and DiT2 in different C3‐ and C4‐type plant species is presented, and we demonstrate that the substrate specificity of DiT2 from the C4‐plant Flaveria bidentis is similar to its counterpart from C3 plants.
Classification and identification of Arabidopsis cell wall mutants using Fourier‐Transform InfraRed (FT‐IR) microspectroscopy
Authors: Mouille Grégory , Robin Stéphane , Lecomte Mannaïg , Pagant Silvère , Höfte Herman 
Published: July 2003

SummaryWe have developed a novel procedure for the rapid classification and identification of Arabidopsis mutants with altered cell wall architecture based on Fourier‐Transform Infrared (FT‐IR) microspectroscopy. FT‐IR transmission spectra were sampled from native 4‐day‐old dark‐grown hypocotyls of 46 mutants and the wild type treated with various drugs. The Mahalanobis distance between mutants, calculated from the spectral information after compression with the Discriminant Variables Selection procedure, was used for α hierarchical cluster analysis. Despite the completely unsupervised nature of the classification procedure, we show that all mutants with cellulose defects appeared in the same cluster. In addition, mutant alleles of similar strength for several unrelated loci were also clustered, which demonstrates the sensitivity of the method to detect a wide array of cell wall defects. Comparing the cellulose‐deficient cluster with the cluster that contained wild‐type controls led to the identification of wave numbers that were diagnostic for altered cellulose content in the context of an intact cell wall. The results show that FT‐IR spectra can be used to identify different classes of mutants and to characterize cell wall changes at a microscopic level in unknown mutants. This procedure significantly accelerates the identification and classification of cell wall mutants, which makes cell wall polysaccharides more accessible to functional genomics approaches.
Development of enhancer trap lines for functional analysis of the rice genome
Authors: Wu Changyin , Li Xiangjun , Yuan Wenya , Chen Guoxing , Kilian Andrzej , Li Juan , Xu Caiguo , Li Xianghua , Zhou Dao‐Xiu , Wang Shiping , Zhang Qifa 
Published: July 2003

SummaryEnhancer trapping has provided a powerful strategy for identifying novel genes and regulatory elements. In this study, we adopted an enhancer trap system, consisting of the GAL4/VP16–UAS elements with GUS as the reporter, to generate a trapping population of rice. Currently, 31 443 independent transformants were obtained from two cultivars using Agrobacterium‐mediated T‐DNA insertion. PCR tests and DNA blot hybridization showed that about 94% of the transformants contained T‐DNA insertions. The transformants carried, on average, two copies of the T‐DNA, and 42% of the transformants had single‐copy insertions. Histochemical assays of approximately 1000 T0 plants revealed various patterns of the reporter gene expression, including expression in only one tissue, and simultaneously in two or more tissues. The expression pattern of the reporter gene in T1 families corresponded well with the T0 plants and segregated in a 3 : 1 Mendelian ratio in majority of the T1 families tested. The frequency of reporter gene expression in the enhancer trap lines was much higher than that in gene trap lines reported previously. Analysis of flanking sequences of T‐DNA insertion sites from about 200 transformants showed that almost all the sequences had homology with the sequences in the rice genome databases. Morphologically conspicuous mutations were observed in about 7.5% of the 2679 T1 families that were field‐tested, and segregation in more than one‐third of the families fit the 3 : 1 ratio. It was concluded that GAL4/VP16–UAS elements provided a useful system for enhancer trap in rice.
Transgenic tobacco plants expressing antisense ferredoxin‐NADP(H) reductase transcripts display increased susceptibility to photo‐oxidative damage
Authors: Palatnik Javier F. , Tognetti Vanesa B. , Poli Hugo O. , Rodríguez Ramiro E. , Blanco Nicolás , Gattuso Martha , Hajirezaei Mohammad‐Reza , Sonnewald Uwe , Valle Estela M. , Carrillo Néstor 
Published: July 2003

SummaryFerredoxin‐NADP(H) reductase (FNR) catalyses the final step of the photosynthetic electron transport in chloroplasts. Using an antisense RNA strategy to reduce expression of this flavoenzyme in transgenic tobacco plants, it has been demonstrated that FNR mediates a rate‐limiting step of photosynthesis under both limiting and saturating light conditions. Here, we show that these FNR‐deficient plants are abnormally prone to photo‐oxidative injury. When grown under autotrophic conditions for 3 weeks, specimens with 20–40% extant reductase undergo leaf bleaching, lipid peroxidation and membrane damage. The magnitude of the effect was proportional to the light intensity and to the extent of FNR depletion, and was accompanied by morphological changes involving accumulation of aberrant plastids with defective thylakoid stacking. Damage was initially confined to chloroplast membranes, whereas Rubisco and other stromal proteins began to decline only after several weeks of autotrophic growth, paralleled by partial recovery of NADPH levels. Exposure of the transgenic plants to moderately high irradiation resulted in rapid loss of photosynthetic capacity and accumulation of singlet oxygen in leaves. The collected results suggest that the extensive photo‐oxidative damage sustained by plants impaired in FNR expression was caused by singlet oxygen building up to toxic levels in these tissues, as a direct consequence of the over‐reduction of the electron transport chain in FNR‐deficient chloroplasts.
A gene encoding an RNase D exonuclease‐like protein is required for post‐transcriptional silencing in Arabidopsis
Authors: Glazov Eugene , Phillips Kenneth , Budziszewski Gregory J. , Meins Frederick , Levin Joshua Z. 
Published: July 2003

SummaryPost‐transcriptional gene silencing (PTGS) and the closely related phenomenon RNA interference (RNAi) result from the initial endonucleolytic cleavage of target mRNAs, which are then presumed to be completely hydrolyzed by exoribonucleases. To date, no plant genes required for PTGS are known to encode exoribonucleases. The Arabidopsis Werner Syndrome‐like exonuclease (WEX) gene encodes an RNase D domain most similar to that in human Werner Syndrome protein (WRN), but lacks the RecQ helicase domain. It is also related to Caenorhabditis elegans mut‐7, which is essential for RNAi, PTGS, and transposon activity. We isolated a loss‐of‐function mutant, wex‐1, that showed greatly reduced expression of WEX mRNA and early flowering. Although wex‐1 did not affect expression of a robust marker for transcriptional gene silencing (TGS), PTGS of a green‐fluorescent‐protein (GFP) reporter gene was blocked in wex‐1 and restored by ectopic expression of WEX, indicating that WEX is required for PTGS but not TGS. Thus, members of the RNase D protein family are required for PTGS in both plants and animals. Interestingly, WEX has been shown to interact with an Arabidopsis RecQ helicase, suggesting that these proteins might comprise a functional equivalent of WRN.
Absence of the Lhcb1 and Lhcb2 proteins of the light‐harvesting complex of photosystem II – effects on photosynthesis, grana stacking and fitness
Authors: Jenny Andersson , Mark Wentworth , Robin G. Walters , Caroline A. Howard , Alexander V. Ruban , Peter Horton , Stefan Jansson 
Published: July 2003

SummaryWe have constructed Arabidopsis thaliana plants that are virtually devoid of the major light‐harvesting complex, LHC II. This was accomplished by introducing the Lhcb2.1 coding region in the antisense orientation into the genome by Agrobacterium‐mediated transformation. Lhcb1 and Lhcb2 were absent, while Lhcb3, a protein present in LHC II associated with photosystem (PS) II, was retained. Plants had a pale green appearance and showed reduced chlorophyll content and an elevated chlorophyll a/b ratio. The content of PS II reaction centres was unchanged on a leaf area basis, but there was evidence for increases in the relative levels of other light harvesting proteins, notably CP26, associated with PS II, and Lhca4, associated with PS I. Electron microscopy showed the presence of grana. Photosynthetic rates at saturating irradiance were the same in wild‐type and antisense plants, but there was a 10–15% reduction in quantum yield that reflected the decrease in light absorption by the leaf. The antisense plants were not able to perform state transitions, and their capacity for non‐photochemical quenching was reduced. There was no difference in growth between wild‐type and antisense plants under controlled climate conditions, but the antisense plants performed worse compared to the wild type in the field, with decreases in seed production of up to 70%.
Target genes for OBP3, a Dof transcription factor, include novel basic helix‐loop‐helix domain proteins inducible by salicylic acid
Authors: Kang Hong‐Gu , Foley Rhonda C. , Oñate‐Sánchez Luis , Lin Chentao , Singh Karam B. 
Published: July 2003

SummaryOverexpression of a salicylic‐acid (SA)‐inducible Arabidopsis DNA binding with one finger (Dof) transcription factor, called OBF‐binding protein 3 (OBP3; AtDof3.6), has previously been shown to result in growth defects. In this study, suppressive subtraction hybridization (SSH) was used to isolate genes induced in an OBP3‐overexpression line and several putative clones, called OBP3‐responsive genes (ORGs), were isolated. The link with the induced expression levels of these genes and OBP3 overexpression was confirmed by analysing additional OBP3‐overexpression lines. ORG1 through ORG4 are novel genes, while ORG5 is an extensin gene, AtExt1. While ORG4 has no similarity with other proteins in the database, ORG1 has weak similarity in different regions of the predicted protein with CDC2 and fibrillin. ORG2 and ORG3 share 80% overall identity in their deduced amino acid sequences and contain a basic helix‐loop‐helix DNA‐binding domain, suggesting that ORG2 and ORG3 may be transcription factors. The expression of the ORG1, ORG2 and ORG3 genes was co‐regulated under all conditions examined including upregulation by SA and downregulation by jasmonic acid (JA). Fifteen OBP3‐silenced lines were generated to further explore the function of OBP3. Although there were no visible phenotypic changes in any of these lines, the expression of ORG1, ORG2 and ORG3 was reduced. Among the ORG genes, ORG1, ORG2 and ORG3 contained the highest number of potential Dof‐binding sites in the promoter region, and their expression was significantly increased within 3 h after induction of OBP3 expression using an inducible promoter system, and closely reflected the expression levels of the exogenous OBP3 protein. The results from the gain‐of‐function and loss‐of‐function experiments suggest that the ORG1, ORG2 and ORG3 genes are direct target genes of OBP3.
The SPA1‐like proteins SPA3 and SPA4 repress photomorphogenesis in the light
Authors: Laubinger Sascha , Hoecker Ute 
Published: July 2003

Summary Suppressor of phyA‐105 (SPA1) is a phytochrome A‐specific signaling intermediate that acts as a light‐dependent repressor of photomorphogenesis in Arabidopsis seedlings. SPA1 is part of a small gene family comprising three genes: SPA1‐related 2 (SPA2), SPA1‐related 3 (SPA3), and SPA1‐related 4 (SPA4). Here, we investigate the functions of SPA3 and SPA4, two very closely related genes coding for proteins with 74% identical amino acids. Seedlings with mutations in SPA3 or SPA4 exhibit enhanced photomorphogenesis in the light, but show no phenotype in darkness. While there are small differences between the effects of spa3 and spa4 mutations, it is apparent that SPA3 and SPA4 function to inhibit light responses in continuous far‐red, red, and blue light. Phytochrome A is necessary for all aspects of the spa4 mutant phenotype, suggesting that SPA4, like SPA1, acts specifically in phytochrome A signaling. Enhanced photoresponsiveness of spa3 mutants is also fully dependent on phytochrome A in far‐red and blue light, but not in red light. Hence, SPA3 function in red light may be dependent on other phytochromes in addition to phytochrome A. Using yeast two‐hybrid and in vitro interaction assays, we further show that SPA3 as well as SPA4 can physically interact with the constitutive repressor of light signaling COP1. Deletion analyses suggest that SPA3 and SPA4, like SPA1, bind to the coiled‐coil domain of COP1. Taken together, our results have identified two new loci coding for negative regulators that may be involved in fine tuning of light responses by interacting with COP1.
Systemic movement of a tobamovirus requires host cell pectin methylesterase
Authors: Chen Min‐Huei , Citovsky Vitaly 
Published: July 2003

SummarySystemic movement of plant viruses through the host vasculature, one of the central events of the infection process, is essential for maximal viral accumulation and development of disease symptoms. The host plant proteins involved in this transport, however, remain unknown. Here, we examined whether or not pectin methylesterase (PME), one of the few cellular proteins known to be involved in local, cell‐to‐cell movement of tobacco mosaic virus (TMV), is also required for the systemic spread of viral infection through the plant vascular system. In a reverse genetics approach, PME levels were reduced in tobacco plants using antisense suppression. The resulting PME antisense plants displayed a significant degree of PME suppression in their vascular tissues but retained the wild‐type pattern of phloem loading and unloading of a fluorescent solute. Systemic transport of TMV in these plants, however, was substantially delayed as compared to the wild‐type tobacco, suggesting a role for PME in TMV systemic infection. Our analysis of virus distribution in the PME antisense plants suggested that TMV systemic movement may be a polar process in which the virions enter and exit the vascular system by two different mechanisms, and it is the viral exit out of the vascular system that involves PME.
Evaluation of in vitro activity of ciclopirox olamine, butenafine HCl and econazole nitrate against dermatophytes, yeasts and bacteria
Authors: Kokjohn Katrina , Bradley Mary , Griffiths Brian , Ghannoum Mahmoud 
Published: July 2003

AbstractBackground  In many instances, a cutaneous fungal infection may exist concomitantly with bacterial involvement. In this study we compared the in vitro activity of three antifungal agents against the dermatophytes, yeasts and bacteria recovered most commonly from cutaneous mycoses and bacterial infections.Methods  Using a microdilution method adapted from the National Committee for Clinical Laboratory Standards (NCCLS), we determined the minimum inhibitory concentrations (MICs) of ciclopirox olamine, econazole nitrate and butenafine HCl against a panel of dermatophyte fungi and yeasts (n = 39) and bacterial isolates (n = 45).Results  All three antifungals demonstrated comparable activity against the dermatophytes tested, with a MIC range of 0.03–0.25 µg/ml for ciclopirox, < 0.001–0.25 µg/ml for econazole and 0.03–0.25 µg/ml for butenafine. For yeasts, ciclopirox showed activity against all isolates, with an MIC range of 0.001–0.25 µg/ml, whereas econazole had a broader range of 0.125–> 0.5 µg/ml. Butenafine displayed limited activity against the yeast Candida albicans and no activity against Malassezia furfur. For the antibacterial activity studies, ciclopirox demonstrated activity against all isolates tested with a range of 0.06–2 µg/ml, while econazole showed activity against Gram‐positive bacteria only, with a MIC range of 0.004–0.25 µg/ml. Butenafine HCl had a limited activity against bacterial isolates tested, showing activity against β‐hemolytic Streptococcus Group A and Corynebacterium only. Neither econazole nitrate nor butenafine HCl demonstrated activity against any of the Gram‐negative strains evaluated in this study.Conclusions   The data suggest that ciclopirox olamine has the broadest in vitro activity, in comparison to econazole and butenafine HCl, against bacteria, yeasts and bacteria. These findings may have implications in the use of these antimycotics in the treatment of mixed cutaneous infections where bacteria or yeasts are present in addition to dermatophytes.
Effect of Nitrogen Rate and Stubble Height on Dry Matter Yield, Crude Protein Content and Crude Protein Yield of a Sorghum–Sudangrass Hybrid[Sorghum bicolor (L.) Moench × Sorghum sudanense (Piper) Stapf.] in the Three‐Cutting System
Authors: Iptas S. , Brohi A. R. 
Published: July 2003

AbstractIn this study, the effects of nitrogen (N) rate (60, 120, 180 and 240 kg N ha−1 applied in three equal dressings at seeding and after the first and second cuttings) and stubble height (7, 14 and 21 cm) on the dry matter (DM) yield, crude protein (CP) content, and CP yield of a sorghum–sudangrass hybrid [Sorghum bicolor (L.) Moench × Sorghum sudanense (Piper) Stapf., cv. Pioneer 988] in the three‐cut system was investigated. The N rate had no significant effect in the first and third cuttings, but in the second cutting DM yields increased significantly with increase in N rate. The highest yield of 9.1 t ha−1 was obtained with 80 kg N ha−1 for the average of 2 years at the second cutting, but no significant difference was found among the 40, 60 and 80 kg N ha−1 rates. CP content and yield were not significantly affected by N rate at the first and third cuttings, but CP content and yield were significantly affected by application of N at the second cutting. Stubble height had a significant effect on CP content at the third cutting. However, it had no significant effect on CP content at the first and second cuttings. Stubble height had a significant effect on the CP yield at the first cutting, but no significant effect on CP yield at the second and third cuttings.