bluuebunny/crossref_metadata_2025 huggingface dataset:
Intrinsic disorder as a mechanism to optimize allosteric coupling in proteins
Authors: Hilser Vincent J. , Thompson E. Brad
Published: May 2007

Transcription factors and other allosteric cell signaling proteins contain a disproportionate number of domains or segments that are intrinsically disordered (ID) under native conditions. In many cases folding of these segments is coupled to binding with one or more of their interaction partners, suggesting that intrinsic disorder plays an important functional role. Despite numerous hypotheses for the role of ID domains in regulation, a mechanistic model has yet to be established that can quantitatively assess the importance of intrinsic disorder for intramolecular site-to-site communication, the hallmark property of allosteric proteins. Here, we present such a model and show that site-to-site allosteric coupling is maximized when intrinsic disorder is present in the domains or segments containing one or both of the coupled binding sites. This result not only explains the prevalence of ID domains in regulatory proteins, it also calls into question the classical mechanical view of energy propagation in proteins, which predicts that site-to-site coupling would be maximized when a well defined pathway of folded structure connects the two sites. Furthermore, in showing that the coupling mechanism conferred by intrinsic disorder is robust and independent of the network of interactions that physically link the coupled sites, unique insights are gained into the energetic ground rules that govern site-to-site communication in all proteins.
Activated Cdc42-associated kinase Ack1 promotes prostate cancer progression via androgen receptor tyrosine phosphorylation
Authors: Mahajan Nupam P. , Liu Yuanbo , Majumder Samarpan , Warren Maria R. , Parker Carol E. , Mohler James L. , Earp H. Shelton , Whang Young E.
Published: May 2007

Activation of the androgen receptor (AR) may play a role in androgen-independent progression of prostate cancer. Multiple mechanisms of AR activation, including stimulation by tyrosine kinases, have been postulated. We and others have recently shown involvement of activated Cdc42-associated tyrosine kinase Ack1 in advanced human prostate cancer. Here we provide the molecular basis for interplay between Ack1 and AR in prostate cancer cells. Activated Ack1 promoted androgen-independent growth of LNCaP and LAPC-4 prostate xenograft tumors, AR recruitment to the androgen-responsive enhancer, and androgen-inducible gene expression in the absence of androgen. Heregulin-stimulated HER2 activation induced Ack1 activation and AR tyrosine phosphorylation. Ack1 knockdown inhibited heregulin-dependent AR tyrosine phosphorylation, AR reporter activity, androgen-stimulated gene expression, and AR recruitment. Ack1 was recruited to the androgen-responsive enhancers after androgen and heregulin stimulation. In 8 of 18 primary androgen-independent prostate tumor samples, tyrosine-phosphorylated AR protein was detected and correlated with the detection of tyrosine-phosphorylated Ack1. Neither was elevated in androgen-dependent tumors or benign prostate samples. Activated Ack1 phosphorylated AR protein at Tyr-267 and Tyr-363, both located within the transactivation domain. Mutation of Tyr-267 completely abrogated and mutation of Tyr-363 reduced Ack1-induced AR reporter activation and recruitment of AR to the androgen-responsive enhancer. Expression of AR point mutants inhibited Ack1-driven xenograft tumor growth. Thus, Ack1 activated by surface signals or oncogenic mechanisms may directly enhance AR transcriptional function and promote androgen-independent progression of prostate cancer. Targeting the Ack1 kinase may be a potential therapeutic strategy in prostate cancer.
Emerging principles of regulatory evolution
Authors: Prud'homme Benjamin , Gompel Nicolas , Carroll Sean B.
Published: May 2007

Understanding the genetic and molecular mechanisms governing the evolution of morphology is a major challenge in biology. Because most animals share a conserved repertoire of body-building and -patterning genes, morphological diversity appears to evolve primarily through changes in the deployment of these genes during development. The complex expression patterns of developmentally regulated genes are typically controlled by numerous independent cis-regulatory elements (CREs). It has been proposed that morphological evolution relies predominantly on changes in the architecture of gene regulatory networks and in particular on functional changes within CREs. Here, we discuss recent experimental studies that support this hypothesis and reveal some unanticipated features of how regulatory evolution occurs. From this growing body of evidence, we identify three key operating principles underlying regulatory evolution, that is, how regulatory evolution: (
i
) uses available genetic components in the form of preexisting and active transcription factors and CREs to generate novelty; (
ii
) minimizes the penalty to overall fitness by introducing discrete changes in gene expression; and (
iii
) allows interactions to arise among any transcription factor and downstream CRE. These principles endow regulatory evolution with a vast creative potential that accounts for both relatively modest morphological differences among closely related species and more profound anatomical divergences among groups at higher taxonomical levels.
Darwin's greatest discovery: Design without designer
Authors: Ayala Francisco J.
Published: May 2007

Darwin's greatest contribution to science is that he completed the Copernican Revolution by drawing out for biology the notion of nature as a system of matter in motion governed by natural laws. With Darwin's discovery of natural selection, the origin and adaptations of organisms were brought into the realm of science. The adaptive features of organisms could now be explained, like the phenomena of the inanimate world, as the result of natural processes, without recourse to an Intelligent Designer. The Copernican and the Darwinian Revolutions may be seen as the two stages of the one Scientific Revolution. They jointly ushered in the beginning of science in the modern sense of the word: explanation through natural laws. Darwin's theory of natural selection accounts for the “design” of organisms, and for their wondrous diversity, as the result of natural processes, the gradual accumulation of spontaneously arisen variations (mutations) sorted out by natural selection. Which characteristics will be selected depends on which variations happen to be present at a given time in a given place. This in turn depends on the random process of mutation as well as on the previous history of the organisms. Mutation and selection have jointly driven the marvelous process that, starting from microscopic organisms, has yielded orchids, birds, and humans. The theory of evolution conveys chance and necessity, randomness and determinism, jointly enmeshed in the stuff of life. This was Darwin's fundamental discovery, that there is a process that is creative, although not conscious.
On the origin and evolutionary diversification of beetle horns
Authors: Emlen Douglas J. , Corley Lavine Laura , Ewen-Campen Ben
Published: May 2007

Many scarab beetles produce rigid projections from the body called horns. The exaggerated sizes of these structures and the staggering diversity of their forms have impressed biologists for centuries. Recent comparative studies using DNA sequence-based phylogenies have begun to reconstruct the historical patterns of beetle horn evolution. At the same time, developmental genetic experiments have begun to elucidate how beetle horns grow and how horn growth is modulated in response to environmental variables, such as nutrition. We bring together these two perspectives to show that they converge on very similar conclusions regarding beetle evolution. Horns do not appear to be difficult structures to gain or lose, and they can diverge both dramatically and rapidly in form. Although much of this work is still preliminary, we use available information to propose a conceptual developmental model for the major trajectories of beetle horn evolution. We illustrate putative mechanisms underlying the evolutionary origin of horns and the evolution of horn location, shape, allometry, and dimorphism.
Insect societies as divided organisms: The complexities of purpose and cross-purpose
Authors: Strassmann Joan E. , Queller David C.
Published: May 2007

Individual organisms are complex in a special way. The organization and function of their parts seem directed toward a purpose: the survival and reproduction of that individual. Groups of organisms are different. They may also be complex, but that is usually because their parts, the individual organisms, are working at cross-purposes. The most obvious exception to this rule is the social insects. Here, the individuals cooperate in complex ways toward the common goal of the success of the colony, even if it means that most of them do not reproduce. Kin selection theory explains how this can evolve. Nonreproductive individuals help in the reproduction of their kin, who share and transmit their genes. Such help is most favored when individuals can give more to their kin than they give up by not reproducing directly. For example, they can remain at their natal site and help defend a valuable resource (“fortress defenders”), or they can ensure that at least one adult survives to care for helpless young (“life insurers”). Although kin selection explains the extensive cooperation and common purpose of social insect colonies, it also predicts a certain amount of cross-purpose and conflict behavior. Kin selection has predicted how workers and queens disagree over sex ratios, how potential queens struggle to be the colony's head, how workers try to produce sons, and how other workers often prevent them. Kin selection analysis of cooperation and conflict in social insects is one of the outstanding achievements of evolutionary theory.
The neoselectionist theory of genome evolution
Authors: Bernardi Giorgio
Published: May 2007

The vertebrate genome is a mosaic of GC-poor and GC-rich isochores, megabase-sized DNA regions of fairly homogeneous base composition that differ in relative amount, gene density, gene expression, replication timing, and recombination frequency. At the emergence of warm-blooded vertebrates, the gene-rich, moderately GC-rich isochores of the cold-blooded ancestors underwent a GC increase. This increase was similar in mammals and birds and was maintained during the evolution of mammalian and avian orders. Neither the GC increase nor its conservation can be accounted for by the random fixation of neutral or nearly neutral single-nucleotide changes (i.e., the vast majority of nucleotide substitutions) or by a biased gene conversion process occurring at random genome locations. Both phenomena can be explained, however, by the neoselectionist theory of genome evolution that is presented here. This theory fully accepts Ohta's nearly neutral view of point mutations but proposes in addition (
i
) that the AT-biased mutational input present in vertebrates pushes some DNA regions below a certain GC threshold; (
ii
) that these lower GC levels cause regional changes in chromatin structure that lead to deleterious effects on replication and transcription; and (
iii
) that the carriers of these changes undergo negative (purifying) selection, the final result being a compositional conservation of the original isochore pattern in the surviving population. Negative selection may also largely explain the GC increase accompanying the emergence of warm-blooded vertebrates. In conclusion, the neoselectionist theory not only provides a solution to the neutralist/selectionist debate but also introduces an epigenomic component in genome evolution.
Two routes to functional adaptation: Tibetan and Andean high-altitude natives
Authors: Beall Cynthia M.
Published: May 2007

Populations native to the Tibetan and Andean Plateaus are descended from colonizers who arrived perhaps 25,000 and 11,000 years ago, respectively. Both have been exposed to the opportunity for natural selection for traits that offset the unavoidable environmental stress of severe lifelong high-altitude hypoxia. This paper presents evidence that Tibetan and Andean high-altitude natives have adapted differently, as indicated by large quantitative differences in numerous physiological traits comprising the oxygen delivery process. These findings suggest the hypothesis that evolutionary processes have tinkered differently on the two founding populations and their descendents, with the result that the two followed different routes to the same functional outcome of successful oxygen delivery, long-term persistence and high function. Assessed on the basis of basal and maximal oxygen consumption, both populations avail themselves of essentially the full range of oxygen-using metabolism as populations at sea level, in contrast with the curtailed range available to visitors at high altitudes. Efforts to identify the genetic bases of these traits have included quantitative genetics, genetic admixture, and candidate gene approaches. These reveal generally more genetic variance in the Tibetan population and more potential for natural selection. There is evidence that natural selection is ongoing in the Tibetan population, where women estimated to have genotypes for high oxygen saturation of hemoglobin (and less physiological stress) have higher offspring survival. Identifying the genetic bases of these traits is crucial to discovering the steps along the Tibetan and Andean routes to functional adaptation.
An experimental test of evolutionary trade-offs during temperature adaptation
Authors: Bennett Albert F. , Lenski Richard E.
Published: May 2007

We used experimental evolution to test directly the important and commonplace evolutionary hypothesis that adaptation, increased fitness within the selective environment, is accompanied by trade-off, a loss of fitness in other nonselective environments. Specifically, we determined whether trade-offs at high temperature generally and necessarily accompany genetic adaptation to low temperature. We measured the relative fitness increment of 24 lineages of the bacterium
Escherichia coli
evolved for 2,000 generations at 20°C and the relative fitness decrement of these lines at 40°C. Trade-offs at the higher temperature were examined for their generality, universality, quantitative relationship, and historical contingency. Considering all 24 lines as a group, a significant decline in fitness was found at 40°C (mean decline = 9.4%), indicating the generality of the trade-off effect. However, in a lineage-by-lineage analysis, only 15 of 24 showed a significant trade-off, and one lineage increased fitness at high temperature. Thus, although general, trade-offs were not universal. Furthermore, there was no quantitative association between the magnitude of adaptive fitness increment at 20°C and fitness decline at 40°C, and no effect of lineages' historical thermal environment on either their improvement at 20°C or the extent of their trade-off at high temperature. We do not yet know the underlying mechanisms responsible for the trade-off, but they are sufficiently prevalent to drive a general effect. However, approximately one-third of the experimental lineages achieved low-temperature adaptation without detectable high-temperature trade-offs; therefore, it cannot be necessary that every change conferring benefit in cold environments has a negative effect on function in warmer environments.
Hepatitis C virus epitope-specific neutralizing antibodies in Igs prepared from human plasma
Authors: Zhang Pei , Wu Charles G. , Mihalik Kathleen , Virata Maria Luisa , Yu Mei-ying W. , Alter Harvey J. , Feinstone Stephen M.
Published: May 2007

Neutralizing antibodies directed against hepatitis C virus (HCV) are present in Igs made from anti-HCV-positive plasma. However, these HCV-specific Igs are largely ineffective
in vivo
. The mechanism for the poor effectiveness is currently unknown. We hypothesize that the presence of nonneutralizing antibodies in HCV-specific Igs interferes with the function of neutralizing antibodies, resulting in the reduction or blockage of their effect. In the present study, we identified at least two epitopes at amino acid residues 412–419 (epitope I) and 434–446 (epitope II), located downstream of the hypervariable region I within the HCV E2 protein. We demonstrated that epitope I, but not epitope II, was implicated in HCV neutralization and that binding of a nonneutralizing antibody to epitope II completely disrupted virus neutralization mediated by antibody binding at epitope I. The dynamic interaction between nonneutralizing and neutralizing antibodies may thus play a key role in determining the outcomes of HCV infection. Further exploration of this interplay should lead to a better understanding of the mechanisms of neutralization and immune escape and may indicate pathways for the manufacture of an effective HCV-specific Ig product for immune prophylaxis of HCV infection.
Depletion of the Nucleolar Protein Nucleostemin Causes G1 Cell Cycle Arrest via the p53 Pathway
Authors: Ma Hanhui , Pederson Thoru
Published: May 2007

Nucleostemin (NS) is a nucleolar protein expressed in adult and embryo-derived stem cells, transformed cell lines, and tumors. NS decreases when proliferating cells exit the cell cycle, but it is unknown how NS is controlled, and how it participates in cell growth regulation. Here, we show that NS is down-regulated by the tumor suppressor p14ARF and that NS knockdown elevates the level of tumor suppressor p53. NS knockdown led to G1 cell cycle arrest in p53-positive cells but not in cells in which p53 was genetically deficient or depleted by small interfering RNA knockdown. These results demonstrate that, in the cells investigated, the level of NS is regulated by p14ARF and the control of the G1/S transition by NS operates in a p53-dependent manner.
Biophoton Imaging: A Nondestructive Method for Assaying R Gene Responses
Authors: Bennett Mark , Mehta Monaz , Grant Murray
Published: May 2007

Plant disease resistance (R) proteins of the nucleotide binding-leucine rich repeat class are responsible for pathogen recognition and activation of defense signaling networks leading to the hypersensitive response (HR). Genetically, R-protein signaling appears to be integrated through a limited set of common downstream components. However, the timing of development of visible HR is unique to individual R proteins. By utilizing the phenomena of ultraweak photon emission from leaves undergoing an incompatible interacttion, a powerful nondestructive and facile assay is described to compare timing of defense responses elicited by different R proteins. We demonstrate that ultraweak photon emission, or “biophoton generation,” is demonstrated to be associated with hypersensitive cell death. Biophoton emission requires an intact R signaling network and increases in cytosolic calcium and nitric oxide, but elevated reactive oxygen species are not necessary. Importantly, the assay is robust and applicable to a range of incompatible interactions in various plant species. The ability to assay R responses nondestructively in real time and a chosen genetic background makes this technique amenable to subtle genetic dissection of plant defense responses.
Recombination Events Generating a Novel Rp1 Race Specificity
Authors: Smith Shavannor M. , Hulbert Scot H.
Published: May 2007

Genes at the maize Rpl rust resistance complex often mispair in meiosis, which allows genes to recombine unequally, creating recombinant haplotypes. Four recombinant haplotypes were identified from progeny of an Rpl-D/Rpl-I heterozygote that conferred a nonparental resistance specificity designated Rpl-I*. Sequence comparisons of paralogs in the recombinant and parental haplotypes demonstrated that all four recombinants were derived from intergenic (between gene) recombination events. The sequence of paralogs in the HRpl-I parental haplotype indicated this haplotype includes 41 or more rpl genes, at least 31 of which are transcribed. The results indicate that most of the novel resistance specificities that have arisen spontaneously at Rp1 are the result of reassortment of existing Rpl genes.
Analysis of Promoter Activity of the Early Nodulin Enod40 in Lotus japonicus
Authors: Grønlund Mette , Roussis Andreas , Flemetakis Emmanouil , Quaedvlieg Nicolette E. M. , Schlaman Helmi R. M. , Umehara Yosuke , Katinakis Panagiotis , Stougaard Jens , Spaink Herman P.
Published: May 2007

Our comparative studies on the promoter (pr) activity of Enod40 in the model legume Lotus japonicus in stably transformed GusA reporter lines and in hairy roots of L. japonicus demonstrate a stringent regulation of the Enod40 promoter in the root cortex and root hairs in response to Nod factors. Interestingly, the L. japonicus Enod40-2 promoter fragment also shows symbiotic activity in the reverse orientation. Deletion analyses of the Glycine max (Gm) Enod40 promoter revealed the presence of a minimal region -185 bp upstream of the transcription start. Stable transgenic L. japonicus reporter lines were used in bioassays to test the effect of different compounds on early symbiotic signaling. The responses of prGmEnod40 reporter lines were compared with the responses of L. japonicus (Lj) reporter lines based on the LjNin promoter. Both reporter lines show very early activity postinoculation in root hairs of the responsive zone of the root and later in the dividing cells of nodule primordia. The LjNin promoter was found to be more responsive than the GmEnod40 promoter to Nod factors and related compounds. The use of prGmEnod40 reporter lines to analyze the effect of nodulin genes on the GmEnod40 promoter activity indicates that LJNIN has a positive effect on the regulation of the Enod40 promoter, whereas the latter is not influenced by ectopic overexpression of its own gene product. In addition to pointing to a difference in the regulation of the two nodulin genes Enod40 and Nin during early time points of symbiosis, the bioassays revealed a difference in the response to the synthetic cytokinin 6-benzylaminopurine (BAP) between alfalfa and clover and L. japonicus. In alfalfa and clover, Enod40 expression was induced upon BAP treatment, whereas this seems not to be the case in L. japonicus; these results correlate with effects at the cellular level because BAP can induce pseudonodules in alfalfa and clover but not in L. japonicus. In conclusion, we demonstrate the applicability of the described L. japonicus reporter lines in analyses of the specificity of compounds related to nodulation as well as for the dissection of the interplay between different nodulin genes.
Sinorhizobium fredii USDA257 Releases a 22-kDa Outer Membrane Protein (Omp22) to the Extracellular Milieu When Grown in Calcium-Limiting Conditions
Authors: Kim Won-Seok , Sun-Hyung Jeong , Park Ro-Dong , Kim Kil-Yong , Krishnan Hari B.
Published: May 2007

Calcium, which regulates a wide variety of cellular functions, plays an important role in Rhizobium-legume interactions. We investigated the effect of calcium on surface appendages of Sinorhizobium fredii USDA257. Cold-field emission scanning electron microscopy observation of USDA257 grown in calcium-limiting conditions revealed cells with unusual shape and size. Transmission electron microscopy observation revealed intact flagella were present only when USDA257 cells were grown in calcium-sufficient conditions. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of flagellar preparations from USDA257 cells grown in calcium-limiting conditions showed the presence of a 22-kDa protein that was absent from cells grown in calcium-sufficient conditions. We have cloned and determined the nucleotide sequence of the gene encoding the 22- kDa protein. After successful expression in Escherichia coli, polyclonal antibodies were raised against the recombinant 22-kDa protein (Omp22). Subcellular fractionation analysis demonstrated that Omp22 was predominantly present in the extracellular fraction. Western blot analysis revealed the presence of immunologically related proteins from diverse rhizobia. Immunocytochemical localization of thin sections of USDA257 cells showed specific labeling of protein A-gold particles on protein inclusions found proximal to the cells. Accumulation of Omp22 was greatly reduced when USDA257 cells were grown in the presence of increasing calcium. Northern blot analysis indicated that calcium was the only divalent cation among those tested that down-regulated omp22 expression. An omp22 mutant was able to grow in calcium-limiting conditions at a rate similar to that of wild-type USDA257. Significantly more nodules were initiated by the omp22 mutant than by the wild-type on soybean cultivar Peking grown in calciumlimiting conditions.
Potato Homologs of Arabidopsis thaliana Genes Functional in Defense Signaling—Identification, Genetic Mapping, and Molecular Cloning
Authors: Pajerowska Karolina M. , Parker Jane E. , Gebhardt Christiane
Published: May 2007

Defense against pests and pathogens is a fundamental process controlled by similar molecular mechanisms in all flowering plants. Using Arabidopsis thaliana as a model, steps of the signal transduction pathways that link pathogen recognition to defense activation have been identified and corresponding genes have been characterized. Defense signaling (DS) genes are functional candidates for controlling natural quantitative variation of resistance to plant pathogens. Nineteen Arabidopsis genes operating in defense signaling cascades were selected. Solanaceae EST (expressed sequence tag) databases were employed to identify the closest homologs of potato (Solanum tuberosum). Sixteen novel DS potato homologs were positioned on the molecular maps. Five DS homologs mapped close to known quantitative resistance loci (QRL) against the oomycete Phytophthora infestans causing late blight and the bacterium Erwinia carotovora subsp. atroseptica causing blackleg of stems and tuber soft rot. The five genes are positional candidates for QRL and are highly sequence related to Arabidopsis genes AtSGT1b, AtPAD4, and AtAOS. Full-length complementary DNA and genomic sequences were obtained for potato genes StSGT1, StPAD4, and StEDS1, the latter being a putative interactor of StPAD4. Our results form the basis for further studies on the contributions of these candidate genes to natural variation of potato disease resistance.
Nuclear DNA Endoreduplication and Expression of the Mitotic Inhibitor Ccs52 Associated to Determinate and Lupinoid Nodule Organogenesis
Authors: González-Sama Alfonso , de la Peña Teodoro Coba , Kevei Zoltan , Mergaert Peter , Lucas M. Mercedes , de Felipe María R. , Kondorosi Eva , Pueyo José J.
Published: May 2007

Lotus japonicus determinate nodules differ greatly from indeterminate nodules in their organogenesis and morphological characteristics, whereas Lupinus albus lupinoid nodules share features of determinate and indeterminate nodules. The mitotic inhibitor Ccs52A is essential for endoreduplication and ploidy-dependent cell enlargement during symbiotic cell differentiation in Medicago truncatula indeterminate nodules. ccs52A homolog genes were isolated from lupin and lotus nodules; the deduced Ccs52A proteins showed high sequence similarity with other Cdh-1-type activators of the anaphase-promoting complex and were grouped with A-type Ccs52 proteins from different plants. In lupin, ccs52A expression was restricted to the earlier stages of nodule development, whereas ccs52A transcripts accumulated in lotus nodule primordia and, to a lesser extent, in mature nodules. Nodule development in Lupinus albus involved a progressive increase in nuclear and cellular size and ploidy level; similarly, Lotus japonicus nodules contained polyploid nuclei and enlarged cells in the infected zone. Nevertheless, in situ hybridization experiments showed the highest ccs52A expression in the inner cortex cells of the lupin nodule primordium, probably associated to the increased size of these cells in mature nodules. In view of our results, Ccs52A-mediated endoreduplication appears to be a universal mechanism required for nodule cell differentiation during the establishment of nitrogen-fixing symbioses.
The Rhizobium leguminosarum biovar trifolii ANU794 Induces Novel Developmental Responses on the Subterranean Clover Cultivar Woogenellup
Authors: Morris Angela Carmen , Djordjevic Michael Anthony
Published: May 2007

The clover-nodulating Rhizobium leguminosarum bv. trifolii ANU794 initiates normal root-nodule development with abnormally low efficiency on the Trifolium subterraneum cv. Woogenellup. The cellular and developmental responses of Woogenellup roots to the site- and dose-defined inoculation of green fluorescent protein (gfp)-labeled cells of ANU843 (nodulation proficient) and ANU794 was investigated using light, fluorescence, and confocal microscopy. Strain ANU794-gfp induced three primordia types and four developmental responses at the inoculation site: true or aberrant nodules (on 5 and 25% of plants, respectively), hybrid structures (20% of plants), or lateral roots (50% of plants). The novel hybrid structures possessed nodule and lateral root-like features and unusual vascular patterning. Strain ANU794-gfp induces lateral root formation by stimulating pericycle cell divisions at all nearby protoxylem poles. Only true nodules induced by ANU794-gfp contained intracellular bacteria. In contrast, strain ANU843-gfp induced nodules only and lateral root formation was suppressed at spot inoculation sites. Primordium types were distinguishable by the emission spectrum characteristics of phenolic UV-absorbing and fluorescent compounds that accumulate in primordium cells. Hybrid primordia contained (at least) two fluorescent cell populations, suggesting that they are chimeric. The results suggest that ANU794 may produce both nodule- and lateral root-generating signals simultaneously.
The Arabidopsis thaliana JASMONATE INSENSITIVE 1 Gene Is Required for Suppression of Salicylic Acid-Dependent Defenses During Infection by Pseudomonas syringae
Authors: Laurie-Berry Neva , Joardar Vinita , Street Ian H. , Kunkel Barbara N.
Published: May 2007

Many plant pathogens suppress antimicrobial defenses using virulence factors that modulate endogenous host defenses. The Pseudomonas syringae phytotoxin coronatine (COR) is believed to promote virulence by acting as a jasmonate analog, because COR-insensitive 1 (coi1) Arabidopsis thaliana and tomato mutants are impaired in jasmonate signaling and exhibit reduced susceptibility to P. syringae. To further investigate the role of jasmonate signaling in disease development, we analyzed several jasmonate-insensitive A. thaliana mutants for susceptibility to P. syringae pv. tomato strain DC3000 and sensitivity to COR. Jasmonate-insensitive1 (jin1) mutants exhibit both reduced susceptibility to P. syringae pv. tomato DC3000 and reduced sensitivity to COR, whereas jasmonate-resistant 1 (jar1) plants exhibit wild-type responses to both COR and P. syringae pv. tomato DC3000. A jin1 jar1 double mutant does not exhibit enhanced jasmonate insensitivity, suggesting that JIN1 functions downstream of jasmonic acid-amino acid conjugates synthesized by JAR1. Reduced disease susceptibility in jin1 mutants is correlated with elevated expression of pathogenesis-related 1(PR-1) and is dependent on accumulation of salicylic acid (SA). We also show that JIN1 is required for normal P. syringae pv. tomato DC3000 symptom development through an SA-independent mechanism. Thus,P. syringae pv. tomatoDC3000 appears to utilize COR to manipulate JIN1-dependent jasmonate signaling both to suppress SA-mediated defenses and to promote symptom development.
Protein Expression Profiles in an Endosymbiotic Cyanobacterium Revealed by a Proteomic Approach
Authors: Ekman Martin , Tollbäck Petter , Klint Johan , Bergman Birgitta
Published: May 2007

Molecular mechanisms behind adaptations in the cyano-bacterium (Nostoc sp.) to a life in endosymbiosis with plants are still not clarified, nor are the interactions between the partners. To get further insights, the proteome of a Nostoc strain, freshly isolated from the symbiotic gland tissue of the angiosperm Gunnera manicata Linden, was analyzed and compared with the proteome of the same strain when free-living. Extracted proteins were separated by two-dimensional gel electrophoresis and were identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry combined with tandem mass spectrometry. Even when the higher percentage of differentiated cells (heterocysts) in symbiosis was compensated for, the majority of the proteins detected in the symbiotic cyanobacteria were present in the free-living counterpart, indicating that most cellular processes were common for both stages. However, differential expression profiling revealed a significant number of proteins to be down-regulated or missing in the symbiotic stage, while others were more abundant or only expressed in symbiosis. The differential protein expression was primarily connected to i) cell envelope-associated processes, including proteins involved in exopolysaccharide synthesis and surface and membrane associated proteins, ii) to changes in growth and metabolic activities (C and N), including upregulation of nitrogenase and proteins involved in the oxidative pentose phosphate pathway and downregu-lation of Calvin cycle enzymes, and iii) to the dark, micro-aerobic conditions offered inside the Gunnera gland cells, including changes in relative phycobiliprotein concentrations. This is the first comprehensive analysis of proteins in the symbiotic state.
MgHog1 Regulates Dimorphism and Pathogenicity in the Fungal Wheat Pathogen Mycosphaerella graminicola
Authors: Mehrabi Rahim , Zwiers Lute-Harm , de Waard Maarten A. , Kema Gert H. J.
Published: May 2007

The dimorphic ascomycete pathogen Mycosphaerella graminicola switches from a yeastlike form to an infectious filamentous form that penetrates the host foliage through stomata. We examined the biological function of the mitogen-activated protein kinase-encoding gene MgHog1 in M. graminicola. Interestingly, MgHog1 mutants were unable to switch to filamentous growth on water agar that mimics the nutritionally poor conditions on the foliar surface and, hence, exclusively developed by a yeastlike budding process. Consequently, due to impaired initiation of infectious germ tubes, as revealed by detailed in planta cytological analyses, the MgHog1 mutants failed to infect wheat leaves. We, therefore, conclude that MgHog1 is a new pathogenicity factor involved in the regulation of dimorphism in M. graminicola. Furthermore, MgHog1 mutants are osmosensitive, resistant to phenylpyrrole and dicarboximide fungicides, and do not melanize.
GmEREBP1 Is a Transcription Factor Activating Defense Genes in Soybean and Arabidopsis
Authors: Mazarei Mitra , Elling Axel A. , Maier Tom R. , Puthoff David P. , Baum Thomas J.
Published: May 2007

Ethylene-responsive element-binding proteins (EREBPs) are plant-specific transcription factors, many of which have been linked to plant defense responses. Conserved EREBP domains bind to the GCC box, a promoter element found in pathogenesis-related (PR) genes. We previously identified an EREBP gene from soybean (GmEREBP1) whose transcript abundance decreased in soybean cyst-nematode-infected roots of a susceptible cultivar, whereas it increased in abundance in infected roots of a resistant cultivar. Here, we report further characterization of this gene. Transient expression analyses showed that GmEREBP1 is localized to the plant nucleus and functions as a transcriptional activator in soybean leaves. Transgenic soybean plants expressing GmEREBP1 activated the expression of the ethylene (ET)-responsive gene PR2 and the ET- and jasmonic acid (JA)-responsive gene PR3, and the salicylic acid (SA)-responsive gene PR1 but not the SA-responsive PR5. Similarly, transgenic Arabidopsis plants expressing GmEREBP1 showed elevated mRNA abundance of the ET-regulated gene PR3 and the ET- and JA-regulated defense-related gene PDF1.2 but not the ET-regulated GST2, and the SA-regulated gene PR1 but not the SA-regulated PR2 and PR5. Transgenic soybean and Arabidopsis plants inoculated with cyst nematodes did not display a significantly altered susceptibility to nematode infection. These results collectively show that GmEREBP1 functions as a transacting inducer of defense gene expression in both soybean and Arabidopsis and mediates the expression of both ET- and JA- and SA-regulated defense-related genes in these plant species.
The Repression of Cell Wall- and Plastid-Related Genes and the Induction of Defense-Related Genes in Rice Plants Infected with Rice dwarf virus
Authors: Shimizu Takumi , Satoh Kouji , Kikuchi Shoshi , Omura Toshihiro
Published: May 2007

An analysis, using microarrays, of gene expression in rice plants infected with Rice dwarf virus revealed significant decreases in levels of expression of genes that are involved in the formation of cell walls, reflecting the stunted growth of diseased plants. The expression of plastid-related genes also was suppressed, as anticipated from the white chlorotic appearance of infected leaves. By contrast, the expression of defense- and stress-related genes was enhanced after viral infection. These results suggest that virus-infected rice plants attempt to survive viral infection and replication by raising the levels of expression of defense- and stress-related genes while suppressing the expression of genes required for the elongation of cells and photosynthesis.
Chalcone Synthase Transcripts Are Detected in Alfalfa Root Hairs Following Inoculation with Wild-TypeRhizobium meliloti
Authors: McKhann Heather I. , Paiva Nancy L. , Dixon Richard A. , Hirsch Ann M.
Published: May 2007

Flavonoids are involved in a number of critical events in the interaction between nitrogen-fixing bacteria and legumes. To get a better understanding of the importance of flavonoids in the earliest stages of the alfalfa-Rhizobium meliloti symbiosis, we followed the expression of two chal-cone synthase (CHS) gene family members as well as of chalcone isomerase (CHI) and isoflavone reductase (IFR) genes. CHS transcripts increased 2 to 4 dpi (days post-inoculation) with wild-type rhizobia, but not after inoculation with the heterologous R. leguminosarum bv. trifolii or with an exopolysaccharide (exo) mutant of R. meliloti. CHS transcripts were detected in the root hairs and epidermal cells of the root hair zone, and infrequently in nodule pri-mordia. Insignificant CHI and IFR mRNA accumulation over control levels was observed in response to rhizobial inoculation. The slight increase in CHS transcript accumulation following wild-type R. meliloti inoculation was correlated with an observed increase in root flavonoid content as well as a change in the nod gene-inducing activity of the root exudate. The nod gene-inducing flavonoids exuded from wild-type rhizobia-inoculated roots were identified as 4′, 7-dihydroxyflavone and 4, 4′ dihydroxy-2′-methoxychalcone. Although there was a slight increase over the uninoculated controls in the level of medicarpin-3-O-glucoside 6″-O-malonate (MGM) in extracts of roots inoculated with rhizobia, IFR transcript accumulation was not significantly elevated over that of the controls. Moreover, no medicarpin aglycone was detected in the inoculated roots. Thus, although inoculation with wild-type rhizobia triggers some of the genes induced during an interaction between a host and a pathogen, the expression of these genes in the Rhizobium-legume interaction is at a very low level, suggesting that rhizobia have evolved a mechanism(s) to avoid triggering the host's defense responses.
High Resolution Mapping of the Indica-Derived Rice Blast Resistance Genes II. Pi-ta2 and Pi-ta and a Consideration of Their Origin
Authors: Rybka Krystyna , Miyamoto Masaru , Ando Ikuo , Saito Akira , Kawasaki Shinji
Published: May 2007

The indica-derived rice blast resistance genes Pi-ta2 and Pi-ta, introgressed into a japonica background, were mapped on the middle of chromosome 12 with restriction fragment length polymorphism (RFLP)/random amplified polymorphic DNA (RAPD) markers and several nearly isogenic lines (NILs). Pi-ta2 cosegregated with XNpb 289 and 196, and was flanked by closely linked markers at 0.3 (ta2-1) and 0.7 cM (XNpb 088, 319/079) on the left (up) side, and at 0.3 (RUBss) and 0.7 cM (XNpb316) on the right (down) side of the map, in F2 analyses of Pi No. 4 × japonica crosses, with 160 recessive homozygotic individuals corresponding to 320 chromosomes. CHEF (contour-clamped hexagonal electric field) Southern blotting suggested that XNpb088 and XNpb239-1, both very close to Pi-ta2, were spanned by a 170-kb band, suggesting about 500 to 750 kb/cM as the physical/genetic distances ratio in the crosses of Pi No. 4 × japonica, which was rather low in recombination ratio, in this region. Pi-ta was mapped to the position overlapping Pi-ta2 by graphical genotype analysis of an NIL with a very narrow introgressed region, Shimokita. This result suggested that these functionally related genes are allelic or at least very closely located, and that they may be derived from a common ancestral gene. However, RFLP analyses of the NILs of these two genes indicated that they are derived from rather different genomes, refuting the report that NILs of Pi-ta share a common donor with those of Pi-ta2. This indicates that the two genes differentiated at a rather ancient time. Two other pedigrees, describing spontaneous recent origins of Pi-ta in japonica, were also refuted and a single indica origin of the gene was indicated.
Requirement of the Movement Protein for Long Distance Spread of Tobacco Mosaic Virus in Grafted Plants
Authors: Arce-Johnson Patricio , Reimann-Philipp Ulrich , Padgett Hal S. , Rivera-Bustamante Rafael , Beachy Roger N.
Published: May 2007

Systemic spread of tobacco mosaic virus (TMV) that lacks a functional movement protein (TMVΔMP) was investigated in grafted tobacco (Nicotiana tabacum) plants. Transgenic plants that express the 30-kDa movement protein (MP) gene (MP) under the control of the rolC (phloem-specific) or pal2 (xylem-specific) promoters were unable to support systemic infection by the mutant virus, while plants that express the MP gene from the cauliflower mosaic virus 35S promoter (35S:MP) led to systemic infection. Doubly grafted plants were constructed in which plants containing the 35S:MP gene were used as root stock and plants carrying various MP constructs constituted the middle scion. The upper scion contained the 35S:MP gene in plants that produce a hypersensitive response when systemically infected by TMV. TMVΔMP moved systemically and produced complete necrosis in the upper scion when expression of MP in the middle scion was under the control of the rolC or 35S promoter, but not when the pal2 promoter was used. When plants expressing a gene encoding a defective MP were used as the middle scion, there was no systemic infection by TMVΔMP, and a delay in systemic infection by wild-type TMV. In grafted plants with middle scions that expressed the TMV 54 kDa gene sequence there was no apparent systemic infection by TMVΔMP in the upper scion. The results obtained indicate that the MP has a role in long distance movement, and support the suggestion that replication is necessary for systemic infection of these grafted plants.
Agrobacterium tumefaciens Transformation of the Radiation Hypersensitive Arabidopsis thaliana Mutants uvh1 and rad5
Authors: Nam Jaesung , Mysore Kirankumar S. , Gelvin Stanton B.
Published: May 2007

The Arabidopsis thaliana mutants uvh1 and rad5, originally identified as radiation hypersensitive, were reported to be deficient in T-DNA integration based on the relative efficiencies of stable transformation and T-DNA transfer. We reassessed these mutants for susceptibility to transformation by Agrobacterium tumefaciens. The mutant rad5 showed a significant reduction in the efficiency of transient as well as stable transformation, compared with its wild-type progenitor. These data indicate that rad5 is blocked at a step in the transformation process prior to T-DNA integration. We additionally found, using both an in vitro root inoculation and an in vivo flower bolt inoculation assay, that the mutant uvh1 is as susceptible to A. tumefaciens-mediated transformation as is its wild-type progenitor, C10.
Opine Catabolic Loci from Agrobacterium Plasmids Confer Chemotaxis to Their Cognate Substrates
Authors: Kim Heenam , Farrand Stephen K.
Published: May 2007

Opines are carbon compounds produced by crown galls and hairy roots induced by Agrobacterium tumefaciens and A. rhizogenes, respectively. These novel condensation products of plant metabolic intermediates are utilized as nutritional sources by the Agrobacterium strains that induced the growths. Thus, opines are thought to favor the propagation of agrobacteria in the tumorsphere. Certain Agrobacterium strains were chemoattracted to opines. The chemotactic activities to octopine, to nopaline, to manno-pine, and to agrocinopines A+B were dependent on the type of the Ti plasmid present in the bacterium. The determinants for chemotaxis to these opines were localized to the regions of the octopine- and nopaline-type Ti plasmids coding for transport and catabolism of that opine. An insertion in accA, which encodes the periplasmic binding protein for agrocinopines A+B, abolished chemotaxis while an insertion in accC, which encodes a component of the transport system, and an insertion in accF, which encodes a function required for agrocinopine catabolism, did not affect chemotaxis to this opine. Thus, transport and catabolism of these opines are not required for the chemo-tactic activity. Analyses of subclones of the acc region confirmed that accA is the only gene required from the Ti plasmid for chemotaxis to agrocinopines A+B.
Membrane Permeabilizing Activity of Pathogenesis-Related Protein Linusitin from Flax Seed
Authors: Anzlovar Sabina , Serra Mauro Dalla , Dermastia Marina , Menestrina Gianfranco
Published: May 2007

Linusitin is a 25-kDa pathogenesis-related (PR) protein of class 5 isolated from flax seeds. It has been proposed that PR-5 proteins exert their antifungal activity by permeabilizing fungal membranes. Using a fluorescent method that has been used for antimicrobial proteins other than PR proteins, we tested this hypothesis. The method is based on the release of the dye calcein from the inside of either small or large unilamellar lipid vesicles. The amount of calcein release induced by the protein was studied as a function of protein and lipid concentration and of membrane composition. All the results could be accounted for with an available statistical model. The model predicts that calcein release from the vesicles is a result of the tetrameric protein aggregation. Whether this aggregate corresponds to a transmembrane pore or to another protein complex that perturbs the membrane remains to be established. The lipid composition of the vesicles affected the permeabilization activity of linusitin. An increase of activity was observed by increasing the content of negatively charged phospholipids in the vesicles, inclusion of sterols into the membrane, or decreasing the pH of the solution. The possible roles of the observed changes in permeabilizing activity in actual plant-fungus interactions are discussed.