Casparian strip diffusion barrier in Arabidopsis is made of a lignin polymer without suberin.

Casparian strips are ring-like cell-wall modifications in the root endodermis of vascular plants. Their presence generates a paracellular barrier, analogous to animal tight junctions, that is thought to be crucial for selective nutrient uptake, exclusion of pathogens, and many other processes. Despite their importance, the chemical nature of Casparian strips has remained a matter of debate, confounding further molecular analysis. Suberin, lignin, lignin-like polymers, or both, have been claimed to make up Casparian strips. Here we show that, in Arabidopsis, suberin is produced much too late to take part in Casparian strip formation. In addition, we have generated plants devoid of any detectable suberin, which still establish functional Casparian strips. In contrast, manipulating lignin biosynthesis abrogates Casparian strip formation. Finally, monolignol feeding and lignin-specific chemical analysis indicates the presence of archetypal lignin in Casparian strips. Our findings establish the chemical nature of the primary root-diffusion barrier in Arabidopsis and enable a mechanistic dissection of the formation of Casparian strips, which are an independent way of generating tight junctions in eukaryotes.

Rationale and first results of developing at-risk (prodromal) criteria for bipolar disorder.

Bipolar affective disorder (BD) is a severe, recurrent and disabling disorder with devastating consequences for individuals, families and society. Although these hazards and costs provide a compelling rationale for development of early detection and early intervention strategies in BD, the development of at-risk criteria for first episode mania is still in an early stage of development. In this paper we review the literature with respect to the clinical, neuroantomical and neuropsychological data, which support this goal. We also describe our recently developed bipolar at-risk criteria (BAR). This criteria comprises the peak age range of the first onset of bipolar disorder, genetic risk, presenting with sub-threshold mania, cyclothymic features or depressive symptoms. An initial pilot evaluation of the BAR criteria in 22 subjects indicated conversion rates to proxies of first-episode mania of 23% within 265 days on average, and high specificity and sensitivity of the criteria. If prospective studies confirm the validity of the BAR criteria, then the criteria would have the potential to open up new avenues of research for indicated prevention in BD and might therefore offer opportunities to ameliorate the severity of, or even prevent BD.

Tissue-specific FLAGELLIN-SENSING 2 (FLS2) expression in roots restores immune responses in Arabidopsis fls2 mutants.

The flagellin receptor of Arabidopsis, At-FLAGELLIN SENSING 2 (FLS2), has become a model for mechanistic and functional studies on plant immune receptors. Responses to flagellin or its active epitope flagellin 22 (flg22) have been extensively studied in Arabidopsis leaves. However, the perception of microbe-associated molecular patterns (MAMPs) and the immune responses in roots are poorly understood. Here, we show that isolated root tissue is able to induce pattern-triggered immunity (PTI) responses upon flg22 perception, in contrast to elf18 (the active epitope of elongation factor thermo unstable (EF-Tu)). Making use of fls2 mutant plants and tissue-specific promoters, we generated transgenic Arabidopsis lines expressing FLS2 only in certain root tissues. This allowed us to study the spatial requirements for flg22 responses in the root. Remarkably, the intensity of the immune responses did not always correlate with the expression level of the FLS2 receptor, but depended on the expressing tissue, supporting the idea that MAMP perception and sensitivity in different tissues contribute to a proper balance of defense responses according to the expected exposure to elicitors. In summary, we conclude that each investigated root tissue is able to perceive flg22 if FLS2 is present and that tissue identity is a major element of MAMP perception in roots.

The cost effectiveness of pharmacological smoking cessation therapies in developing countries: a case study in the Seychelles

OBJECTIVE: To examine the incremental cost effectiveness of the five first line pharmacological smoking cessation therapies in the Seychelles and other developing countries. DESIGN: A Markov chain cohort simulation. SUBJECTS: Two simulated cohorts of smokers: (1) a reference cohort given physician counselling only; (2) a treatment cohort given counselling plus cessation therapy. INTERVENTION: Addition of each of the five pharmacological cessation therapies to physician provided smoking cessation counselling. MAIN OUTCOME MEASURES: Cost per life-year saved (LYS) associated with the five pharmacotherapies. Effectiveness expressed as odds ratios for quitting associated with pharmacotherapies. Costs based on the additional physician time required and retail prices of the medications. RESULTS: Based on prices for currently available generic medications on the global market, the incremental cost per LYS for a 45 year old in the Seychelles was 599 US dollars for gum and 227 dollars for bupropion. Assuming US treatment prices as a conservative estimate, the incremental cost per LYS was significantly higher, though still favourable in comparison to other common medical interventions: 3712 dollars for nicotine gum, 1982 dollars for nicotine patch, 4597 dollars for nicotine spray, 4291 dollars for nicotine inhaler, and 1324 dollars for bupropion. Cost per LYS increased significantly upon application of higher discount rates, which may be used to reflect relatively high opportunity costs for health expenditures in developing countries with highly constrained resources and high overall mortality. CONCLUSION: Pharmacological cessation therapy can be highly cost effective as compared to other common medical interventions in low mortality, middle income countries, particularly if medications can be procured at low prices.

Measuring the diurnal pattern of leaf hyponasty and growth in Arabidopsis - a novel phenotyping approach using laser scanning

Plants forming a rosette during their juvenile growth phase, such as Arabidopsis thaliana (L.) Heynh., are able to adjust the size, position and orientation of their leaves. These growth responses are under the control of the plants circadian clock and follow a characteristic diurnal rhythm. For instance, increased leaf elongation and hyponasty - defined here as the increase in leaf elevation angle - can be observed when plants are shaded. Shading can either be caused by a decrease in the fluence rate of photosynthetically active radiation (direct shade) or a decrease in the fluence rate of red compared with far-red radiation (neighbour detection). In this paper we report on a phenotyping approach based on laser scanning to measure the diurnal pattern of leaf hyponasty and increase in rosette size. In short days, leaves showed constitutively increased leaf elevation angles compared with long days, but the overall diurnal pattern and the magnitude of up and downward leaf movement was independent of daylength. Shade treatment led to elevated leaf angles during the first day of application, but did not affect the magnitude of up and downward leaf movement in the following day. Using our phenotyping device, individual plants can be non-invasively monitored during several days under different light conditions. Hence, it represents a proper tool to phenotype light- and circadian clock-mediated growth responses in order to better understand the underlying regulatory genetic network.

The Th2 lymphoproliferation developing in LatY136F mutant mice triggers polyclonal B cell activation and systemic autoimmunity.

Lat(Y136F) knock-in mice harbor a point mutation in Tyr(136) of the linker for activation of T cells and show accumulation of Th2 effector cells and IgG1 and IgE hypergammaglobulinemia. B cell activation is not a direct effect of the mutation on B cells since in the absence of T cells, mutant B cells do not show an activated phenotype. After adoptive transfer of linker for activation of T cell mutant T cells into wild-type, T cell-deficient recipients, recipient B cells become activated. We show in vivo and in vitro that the Lat(Y136F) mutation promotes T cell-dependent B cell activation leading to germinal center, memory, and plasma cell formation even in an MHC class II-independent manner. All the plasma and memory B cell populations found in physiological T cell-dependent B cell responses are found. Characterization of the abundant plasmablasts found in secondary lymphoid organs of Lat(Y136F) mice revealed the presence of a previously uncharacterized CD93-expressing subpopulation, whose presence was confirmed in wild-type mice after immunization. In Lat(Y136F) mice, B cell activation was polyclonal and not Ag-driven because the increase in serum IgG1 and IgE concentrations involved Abs and autoantibodies with different specificities equally. Although the noncomplement-fixing IgG1 and IgE are the only isotypes significantly increased in Lat(Y136F) serum, we observed early-onset systemic autoimmunity with nephritis showing IgE autoantibody deposits and severe proteinuria. These results show that Th2 cells developing in Lat(Y136F) mice can trigger polyclonal B cell activation and thereby lead to systemic autoimmune disease.

Coordination between zinc and phosphate homeostasis involves the transcription factor PHR1, the phosphate exporter PHO1, and its homologue PHO1;H3 in Arabidopsis.

Interactions between zinc (Zn) and phosphate (Pi) nutrition in plants have long been recognized, but little information is available on their molecular bases and biological significance. This work aimed at examining the effects of Zn deficiency on Pi accumulation in Arabidopsis thaliana and uncovering genes involved in the Zn-Pi synergy. Wild-type plants as well as mutants affected in Pi signalling and transport genes, namely the transcription factor PHR1, the E2-conjugase PHO2, and the Pi exporter PHO1, were examined. Zn deficiency caused an increase in shoot Pi content in the wild type as well as in the pho2 mutant, but not in the phr1 or pho1 mutants. This indicated that PHR1 and PHO1 participate in the coregulation of Zn and Pi homeostasis. Zn deprivation had a very limited effect on transcript levels of Pi-starvation-responsive genes such as AT4, IPS1, and microRNA399, or on of members of the high-affinity Pi transporter family PHT1. Interestingly, one of the PHO1 homologues, PHO1;H3, was upregulated in response to Zn deficiency. The expression pattern of PHO1 and PHO1;H3 were similar, both being expressed in cells of the root vascular cylinder and both localized to the Golgi when expressed transiently in tobacco cells. When grown in Zn-free medium, pho1;h3 mutant plants displayed higher Pi contents in the shoots than wild-type plants. This was, however, not observed in a pho1 pho1;h3 double mutant, suggesting that PHO1;H3 restricts root-to-shoot Pi transfer requiring PHO1 function for Pi homeostasis in response to Zn deficiency.

Insect eggs induce a systemic acquired resistance in Arabidopsis.

Although they constitute an inert stage of the insect's life, eggs trigger plant defences that lead to egg mortality or attraction of egg parasitoids. We recently found that salicylic acid (SA) accumulates in response to oviposition by the Large White butterfly Pieris brassicae, both in local and systemic leaves, and that plants activate a response that is similar to the recognition of pathogen-associated molecular patterns (PAMPs), which are involved in PAMP-triggered immunity (PTI). Here we discovered that natural oviposition by P. brassicae or treatment with egg extract inhibit growth of different Pseudomonas syringae strains in Arabidopsis through the activation of a systemic acquired resistance (SAR). This egg-induced SAR involves the metabolic SAR signal pipecolic acid, depends on ALD1 and FMO1, and is accompanied by a stronger induction of defence genes upon secondary infection. Although P. brassicae larvae showed a reduced performance when feeding on Pseudomonas syringae-infected plants, this effect was less pronounced when infected plants had been previously oviposited. Altogether, our results indicate that egg-induced SAR might have evolved as a strategy to prevent the detrimental effect of bacterial pathogens on feeding larvae.

Nonenzymatic lipid peroxidation reprograms gene expression and activates defense markers in Arabidopsis tocopherol-deficient mutants.

Tocopherols (vitamin E) are lipophilic antioxidants that are synthesized by all plants and are particularly abundant in seeds. Two tocopherol-deficient mutant loci in Arabidopsis thaliana were used to examine the functions of tocopherols in seedlings: vitamin e1 (vte1), which accumulates the pathway intermediate 2,3-dimethyl-5-phytyl-1,4-benzoquinone (DMPBQ); and vte2, which lacks all tocopherols and pathway intermediates. Only vte2 displayed severe seedling growth defects, which corresponded with massively increased levels of the major classes of nonenzymatic lipid peroxidation products: hydroxy fatty acids, malondialdehyde, and phytoprostanes. In the absence of pathogens, the phytoalexin camalexin accumulated in vte2 seedlings to levels 100-fold higher than in wild-type or vte1 seedlings. Similarly, gene expression profiling in wild-type, vte1, and vte2 seedlings indicated that increased levels of nonenzymatic lipid peroxidation in vte2 corresponded to increased expression of many defense-related genes, which were not induced in vte1. Both biochemical and transcriptional analyses of vte2 seedlings indicate that nonenzymatic lipid peroxidation plays a significant role in modulating plant defense responses. Together, these results establish that tocopherols in wild-type plants or DMPBQ in vte1 plants limit nonenzymatic lipid peroxidation during germination and early seedling development, thereby preventing the inappropriate activation of transcriptional and biochemical defense responses.

Developing psychosis and its risk States through the lens of schizotypy.

Starting from the early descriptions of Kraepelin and Bleuler, the construct of schizotypy was developed from observations of aberrations in nonpsychotic family members of schizophrenia patients. In contemporary diagnostic manuals, the positive symptoms of schizotypal personality disorder were included in the ultra high-risk (UHR) criteria 20 years ago, and nowadays are broadly employed in clinical early detection of psychosis. The schizotypy construct, now dissociated from strict familial risk, also informed research on the liability to develop any psychotic disorder, and in particular schizophrenia-spectrum disorders, even outside clinical settings. Against the historical background of schizotypy it is surprising that evidence from longitudinal studies linking schizotypy, UHR, and conversion to psychosis has only recently emerged; and it still remains unclear how schizotypy may be positioned in high-risk research. Following a comprehensive literature search, we review 18 prospective studies on 15 samples examining the evidence for a link between trait schizotypy and conversion to psychosis in 4 different types of samples: general population, clinical risk samples according to UHR and/or basic symptom criteria, genetic (familial) risk, and clinical samples at-risk for a nonpsychotic schizophrenia-spectrum diagnosis. These prospective studies underline the value of schizotypy in high-risk research, but also point to the lack of evidence needed to better define the position of the construct of schizotypy within a developmental psychopathology perspective of emerging psychosis and schizophrenia-spectrum disorders.

MR connectomics: a conceptual framework for studying the developing brain.

THE COMBINATION OF ADVANCED NEUROIMAGING TECHNIQUES AND MAJOR DEVELOPMENTS IN COMPLEX NETWORK SCIENCE, HAVE GIVEN BIRTH TO A NEW FRAMEWORK FOR STUDYING THE BRAIN: "connectomics." This framework provides the ability to describe and study the brain as a dynamic network and to explore how the coordination and integration of information processing may occur. In recent years this framework has been used to investigate the developing brain and has shed light on many dynamic changes occurring from infancy through adulthood. The aim of this article is to review this work and to discuss what we have learned from it. We will also use this body of work to highlight key technical aspects that are necessary in general for successful connectome analysis using today's advanced neuroimaging techniques. We look to identify current limitations of such approaches, what can be improved, and how these points generalize to other topics in connectome research.

A hyperactive quantitative trait locus allele of Arabidopsis BRX contributes to natural variation in root growth vigor.

Quantitative trait loci analysis of natural Arabidopsis thaliana accessions is increasingly exploited for gene isolation. However, to date this has mostly revealed deleterious mutations. Among them, a loss-of-function allele identified the root growth regulator BREVIS RADIX (BRX). Here we present evidence that BRX and the paralogous BRX-LIKE (BRXL) genes are under selective constraint in monocotyledons as well as dicotyledons. Unexpectedly, however, whereas none of the Arabidopsis orthologs except AtBRXL1 could complement brx null mutants when expressed constitutively, nearly all monocotyledon BRXLs tested could. Thus, BRXL proteins seem to be more diversified in dicotyledons than in monocotyledons. This functional diversification was correlated with accelerated rates of sequence divergence in the N-terminal regions. Population genetic analyses of 30 haplotypes are suggestive of an adaptive role of AtBRX and AtBRXL1. In two accessions, Lc-0 and Lov-5, seven amino acids are deleted in the variable region between the highly conserved C-terminal, so-called BRX domains. Genotyping of 42 additional accessions also found this deletion in Kz-1, Pu2-7, and Ws-0. In segregating recombinant inbred lines, the Lc-0 allele (AtBRX(Lc-0)) conferred significantly enhanced root growth. Moreover, when constitutively expressed in the same regulatory context, AtBRX(Lc-0) complemented brx mutants more efficiently than an allele without deletion. The same was observed for AtBRXL1, which compared with AtBRX carries a 13 amino acid deletion that encompasses the deletion found in AtBRX(Lc-0). Thus, the AtBRX(Lc-0) allele seems to contribute to natural variation in root growth vigor and provides a rare example of an experimentally confirmed, hyperactive allelic variant.

Differentially phased leaf growth and movements in Arabidopsis depend on coordinated circadian and light regulation.

In contrast to vastly studied hypocotyl growth, little is known about diel regulation of leaf growth and its coordination with movements such as changes in leaf elevation angle (hyponasty). We developed a 3D live-leaf growth analysis system enabling simultaneous monitoring of growth and movements. Leaf growth is maximal several hours after dawn, requires light, and is regulated by daylength, suggesting coupling between growth and metabolism. We identify both blade and petiole positioning as important components of leaf movements in Arabidopsis thaliana and reveal a temporal delay between growth and movements. In hypocotyls, the combination of circadian expression of PHYTOCHROME INTERACTING FACTOR4 (PIF4) and PIF5 and their light-regulated protein stability drives rhythmic hypocotyl elongation with peak growth at dawn. We find that PIF4 and PIF5 are not essential to sustain rhythmic leaf growth but influence their amplitude. Furthermore, EARLY FLOWERING3, a member of the evening complex (EC), is required to maintain the correct phase between growth and movement. Our study shows that the mechanisms underlying rhythmic hypocotyl and leaf growth differ. Moreover, we reveal the temporal relationship between leaf elongation and movements and demonstrate the importance of the EC for the coordination of these phenotypic traits.

Adenosine A1 receptor activation is arrhythmogenic in the developing heart through NADPH oxidase/ERK- and PLC/PKC-dependent mechanisms.

Whether adenosine, a crucial regulator of the developing cardiovascular system, can provoke arrhythmias in the embryonic/fetal heart remains controversial. Here, we aimed to establish a mechanistic basis of how an adenosinergic stimulation alters function of the developing heart. Spontaneously beating hearts or dissected atria and ventricle obtained from 4-day-old chick embryos were exposed to adenosine or specific agonists of the receptors A(1)AR (CCPA), A(2A)AR (CGS-21680) and A(3)AR (IB-MECA). Expression of the receptors was determined by quantitative PCR. The functional consequences of blockade of NADPH oxidase, extracellular signal-regulated kinase (ERK), phospholipase C (PLC), protein kinase C (PKC) and L-type calcium channel (LCC) in combination with adenosine or CCPA, were investigated in vitro by electrocardiography. Furthermore, the time-course of ERK phosphorylation was determined by western blotting. Expression of A(1)AR, A(2A)AR and A(2B)AR was higher in atria than in ventricle while A(3)AR was equally expressed. Adenosine (100μM) triggered transient atrial ectopy and second degree atrio-ventricular blocks (AVB) whereas CCPA induced mainly Mobitz type I AVB. Atrial rhythm and atrio-ventricular propagation fully recovered after 60min. These arrhythmias were prevented by the specific A(1)AR antagonist DPCPX. Adenosine and CCPA transiently increased ERK phosphorylation and induced arrhythmias in isolated atria but not in ventricle. By contrast, A(2A)AR and A(3)AR agonists had no effect. Interestingly, the proarrhythmic effect of A(1)AR stimulation was markedly reduced by inhibition of NADPH oxidase, ERK, PLC, PKC or LCC. Moreover, NADPH oxidase inhibition or antioxidant MPG prevented both A(1)AR-mediated arrhythmias and ERK phosphorylation. These results suggest that pacemaking and conduction disturbances are induced via A(1)AR through concomitant stimulation of NADPH oxidase and PLC, followed by downstream activation of ERK and PKC with LCC as possible target.

Natural genetic variation in Arabidopsis: tools, traits and prospects for evolutionary ecology.

BACKGROUND: The model plant Arabidopsis thaliana (Arabidopsis) shows a wide range of genetic and trait variation among wild accessions. Because of its unparalleled biological and genomic resources, the potential of Arabidopsis for molecular genetic analysis of this natural variation has increased dramatically in recent years. SCOPE: Advanced genomics has accelerated molecular phylogenetic analysis and gene identification by quantitative trait loci (QTL) mapping and/or association mapping in Arabidopsis. In particular, QTL mapping utilizing natural accessions is now becoming a major strategy of gene isolation, offering an alternative to artificial mutant lines. Furthermore, the genomic information is used by researchers to uncover the signature of natural selection acting on the genes that contribute to phenotypic variation. The evolutionary significance of such genes has been evaluated in traits such as disease resistance and flowering time. However, although molecular hallmarks of selection have been found for the genes in question, a corresponding ecological scenario of adaptive evolution has been difficult to prove. Ecological strategies, including reciprocal transplant experiments and competition experiments, and utilizing near-isogenic lines of alleles of interest will be a powerful tool to measure the relative fitness of phenotypic and/or allelic variants. CONCLUSIONS: As the plant model organism, Arabidopsis provides a wealth of molecular background information for evolutionary genetics. Because genetic diversity between and within Arabidopsis populations is much higher than anticipated, combining this background information with ecological approaches might well establish Arabidopsis as a model organism for plant evolutionary ecology.