Water-soluble chlorophyll protein (WSCP) of Arabidopsis is expressed in the gynoecium and developing silique

Water-soluble chlorophyll protein (WSCP) has been found in many Brassicaceae, most often in leaves. In many cases, its expression is stress-induced, therefore, it is thought to be involved in some stress response. In this work, recombinant WSCP from Arabidopsis thaliana (AtWSCP) is found to form chlorophyll-protein complexes in vitro that share many properties with recombinant or native WSCP from Brassica oleracea, BoWSCP, including an unusual heat resistance up to 100°C in aqueous solution. A polyclonal antibody raised against the recombinant apoprotein is used to identify plant tissues expressing AtWSCP. The only plant organs containing significant amounts of AtWSCP are the gynoecium in open flowers and the septum of developing siliques, specifically the transmission tract. In fully grown but still green siliques, the protein has almost disappeared. Possible implications for AtWSCP functions are discussed.     

DNA fingerprinting validates seed dispersal curves from observational studies in the neotropical legume parkia

Background

Determining the distances over which seeds are dispersed is a crucial component for examining spatial patterns of seed dispersal and their consequences for plant reproductive success and population structure. However, following the fate of individual seeds after removal from the source tree till deposition at a distant place is generally extremely difficult. Here we provide a comparison of observationally and genetically determined seed dispersal distances and dispersal curves in a Neotropical animal-plant system.

Methodology/Principal Findings

In a field study on the dispersal of seeds of three Parkia (Fabaceae) species by two Neotropical primate species, Saguinus fuscicollis and Saguinus mystax, in Peruvian Amazonia, we observationally determined dispersal distances. These dispersal distances were then validated through DNA fingerprinting, by matching DNA from the maternally derived seed coat to DNA from potential source trees. We found that dispersal distances are strongly right-skewed, and that distributions obtained through observational and genetic methods and fitted distributions do not differ significantly from each other.

Conclusions/Significance

Our study showed that seed dispersal distances can be reliably estimated through observational methods when a strict criterion for inclusion of seeds is observed. Furthermore, dispersal distances produced by the two primate species indicated that these primates fulfil one of the criteria for efficient seed dispersers. Finally, our study demonstrated that DNA extraction methods so far employed for temperate plant species can be successfully used for hard-seeded tropical plants.     

C-terminal fluorescent labeling impairs functionality of DNA mismatch repair proteins

The human DNA mismatch repair (MMR) process is crucial to maintain the integrity of the genome and requires many different proteins which interact perfectly and coordinated. Germline mutations in MMR genes are responsible for the development of the hereditary form of colorectal cancer called Lynch syndrome. Various mutations mainly in two MMR proteins, MLH1 and MSH2, have been identified so far, whereas 55% are detected within MLH1, the essential component of the heterodimer MutLα (MLH1 and PMS2). Most of those MLH1 variants are pathogenic but the relevance of missense mutations often remains unclear. Many different recombinant systems are applied to filter out disease-associated proteins whereby fluorescent tagged proteins are frequently used. However, dye labeling might have deleterious effects on MutLα's functionality. Therefore, we analyzed the consequences of N- and C-terminal fluorescent labeling on expression level, cellular localization and MMR activity of MutLα. Besides significant influence of GFP- or Red-fusion on protein expression we detected incorrect shuttling of single expressed C-terminal GFP-tagged PMS2 into the nucleus and found that C-terminal dye labeling impaired MMR function of MutLα. In contrast, N-terminal tagged MutLαs retained correct functionality and can be recommended both for the analysis of cellular localization and MMR efficiency.     

Interaction between IRF6 and TGFA Genes Contribute to the Risk of Nonsyndromic Cleft Lip/Palate

Previous evidence from tooth agenesis studies suggested IRF6 and TGFA interact. Since tooth agenesis is commonly found in individuals with cleft lip/palate (CL/P), we used four large cohorts to evaluate if IRF6 and TGFA interaction contributes to CL/P. Markers within and flanking IRF6 and TGFA genes were tested using Taqman or SYBR green chemistries for case-control analyses in 1,000 Brazilian individuals. We looked for evidence of gene-gene interaction between IRF6 and TGFA by testing if markers associated with CL/P were overtransmitted together in the case-control Brazilian dataset and in the additional family datasets. Genotypes for an additional 142 case-parent trios from South America drawn from the Latin American Collaborative Study of Congenital Malformations (ECLAMC), 154 cases from Latvia, and 8,717 individuals from several cohorts were available for replication of tests for interaction. Tgfa and Irf6 expression at critical stages during palatogenesis was analyzed in wild type and Irf6 knockout mice. Markers in and near IRF6 and TGFA were associated with CL/P in the Brazilian cohort (p<10⁻⁶). IRF6 was also associated with cleft palate (CP) with impaction of permanent teeth (p<10⁻⁶). Statistical evidence of interaction between IRF6 and TGFA was found in all data sets (p = 0.013 for Brazilians; p = 0.046 for ECLAMC; p = 10⁻⁶ for Latvians, and p = 0.003 for the 8,717 individuals). Tgfa was not expressed in the palatal tissues of Irf6 knockout mice. IRF6 and TGFA contribute to subsets of CL/P with specific dental anomalies. Moreover, this potential IRF6-TGFA interaction may account for as much as 1% to 10% of CL/P cases. The Irf6-knockout model further supports the evidence of IRF6-TGFA interaction found in humans.     

BCL3 gene role in facial morphology

BACKGROUND: Cleft lip (CL) with or without palate (CLP) and isolated cleft palate (CP) are etiologically complex diseases with interactions among various environmental and genetic factors. The aim of the current study was to identify association with genetic markers and phenotypic craniofacial data in patients with CL/CLP/CP parents. METHODS: Posteroanterior and lateral digital radiographs of the cranium were obtained from 74 parents of patients with CL/CLP/CP. One hundred seventy‐three patients with CL/CLP/CP and 190 controls were enrolled in the study for the association test. Five genetic markers of the IRF6 gene and 14 markers of the 19q13 locus were genotyped. Linear regression analysis was performed for the relationship of cephalometric measurements with genotype data adjusted for age, gender, and cleft type. Chi‐square and transmission disequilibrium tests were performed to evaluate differences in alleles of the BCL3 gene. Positive findings were replicated in an independent sample (n = 95) of patients with CL/CLP/CP parents. RESULTS: Genetic markers of the BCL3 gene at 19q13, rs7257231, and rs1979377 in the familial association test and rs10401176 in the case‐control association test, were associated with craniofacial phenotype. Carriers of BCL3 allele rs7257231T had longer posterior cranial bases than noncarriers (p_adjusted = 0.0028), and in the familial‐based association test showed the statistically strongest relationship (p_adjusted = 0.05) to phenotype. Relation of rs7257231 to facial formation was confirmed in the replication group (p = 0.0024). CONCLUSIONS: The results indicate that BCL3, which has functions related to cell adhesion and whose downregulation can cause disruption of ectodermal development, is likely to be important in facial formation. Birth Defects Research (Part A), 2012. © 2012 Wiley Periodicals, Inc.     

Membrane Region M2C2 in Subunit KtrB of the K+ Uptake System KtrAB from Vibrio alginolyticus Forms a Flexible Gate Controlling K+ Flux: AN ELECTRON PARAMAGNETIC RESONANCE STUDY*

Transmembrane stretch M_2C from the bacterial K⁺-translocating protein KtrB is unusually long. In its middle part, termed M_2C2, it contains several small and polar amino acids. This region is flanked by the two α-helices M_2C1 and M_2C3 and may form a flexible gate at the cytoplasmic side of the membrane controlling K⁺ translocation. In this study, we provide experimental evidence for this notion by using continuous wave and pulse EPR measurements of single and double spin-labeled cysteine variants of KtrB. Most of the spin-labeled residues in M_2C2 were shown to be immobile, pointing to a compact structure. However, the high polarity revealed for the microenvironment of residue positions 317, 318, and 327 indicated the existence of a water-accessible cavity. Upon the addition of K⁺ ions, M_2C2 residue Thr-318R1 (R1 indicates the bound spin label) moved with respect to M_2B residue Asp-222R1 and M_2C3 residue Val-331R1 but not with respect to M_2C1 residue Met-311R1. Based on distances determined between spin-labeled residues of double-labeled variants of KtrB in the presence and absence of K⁺ ions, structural models of the open and closed conformations were developed.     

Voxel-Based Morphometry in Women with Borderline Personality Disorder with and without Comorbid Posttraumatic Stress Disorder

Patients with Borderline Personality Disorder (BPD) showed reduced volume of amygdala and hippocampus, but similar findings are evident in Posttraumatic Stress Disorder (PTSD). Applying voxel-based morphometry (VBM) in a larger cohort of patients with BPD, we sought to extend earlier findings of volume abnormalities in limbic regions and to evaluate the influence of co-occurring PTSD in BPD patients. We used voxel-based morphometry to study gray matter volume (GMV) in 60 healthy controls (HC) and 60 patients with BPD. Subgroup analyses on 53 patients concerning the role of co-occurring PTSD were conducted. Additionally, regression analyses were calculated to assess the relation between borderline symptom severity as well as dissociative experiences and GMV. Differences in local GMV between patients with BPD and HC were observed in the amygdale and hippocampus as well as in the fusiform and cingulate gyrus. Co-occurring PTSD was accompanied by increased GMV in the superior temporal gyrus and dorsolateral prefrontal cortex. Independent of co-occurring PTSD, severity of BPD symptoms predicted smaller GMV in the amygdala and dorsal ACC. Dissociation was positively related to GMV in the middle temporal gyrus. We could replicate earlier findings of diminished limbic GMV in patients with BPD and additionally show that patients with co-morbid PTSD feature increased GMV in prefrontal regions associated with cognitive control.     

The Effect of Hydraulic Loading Rate and Influent Source on the Binding Capacity of Phosphorus Filters

Sorption by active filter media can be a convenient option for phosphorus (P) removal and recovery from wastewater for on-site treatment systems. There is a need for a robust laboratory method for the investigation of filter materials to enable a reliable estimation of their longevity. The objectives of this study were to (1) investigate and (2) quantify the effect of hydraulic loading rate and influent source (secondary wastewater and synthetic phosphate solution) on P binding capacity determined in laboratory column tests and (3) to study how much time is needed for the P to react with the filter material (reaction time). To study the effects of these factors, a 2² factorial experiment with 11 filter columns was performed. The reaction time was studied in a batch experiment. Both factors significantly (α = 0.05) affected the P binding capacity negatively, but the interaction of the two factors was not significant. Increasing the loading rate from 100 to 1200 L m⁻² d⁻¹ decreased P binding capacity from 1.152 to 0.070 g kg⁻¹ for wastewater filters and from 1.382 to 0.300 g kg⁻¹ for phosphate solution filters. At a loading rate of 100 L m⁻² d⁻¹, the average P binding capacity of wastewater filters was 1.152 g kg⁻¹ as opposed to 1.382 g kg⁻¹ for phosphate solution filters. Therefore, influent source or hydraulic loading rate should be carefully controlled in the laboratory. When phosphate solution and wastewater were used, the reaction times for the filters to remove P were determined to be 5 and 15 minutes, respectively, suggesting that a short residence time is required. However, breakthrough in this study occurred unexpectedly quickly, implying that more time is needed for the P that has reacted to be physically retained in the filter.