Changes In Phenotypic Characteristics Research Articles

Chrysanthemum morifolium β-carotene hydroxylase overexpression promotes Arabidopsis thaliana tolerance to high light stress

The β-carotene hydroxylase gene (BCH) regulates zeaxanthin production in response to high light levels ro protect Chrysanthemum morifolium plants against light-induced damage. In this study, the Chrysanthemum morifolium CmBCH1 and CmBCH2 genes were cloned and their functional importance was assessed by overexpressing them in Arabidopsis thaliana. These transgenic plants were evaluated for gene-related changes in phenotypic characteristics, photosynthetic activity, fluorescence properties, carotenoid biosynthesis, aboveground/belowground biomass, pigment content, and the expression of light-regulated genes under conditions of high light stress relative to wild-type (WT) plants. When exposed to high light stress, WT A. thaliana leaves turned yellow and the overall biomass was reduced compared to that of the transgenic plants. WT plants exposed to high light stress also exhibited significant reductions in the net photosynthetic rate, stomatal conductance, Fv/Fm, qP, and ETR, whereas these changes were not observed in the transgenic CmBCH1 and CmBCH2 plants. Lutein and zaxanthin levels were significantly increased in the transgenic CmBCH1 and CmBCH2 lines, with progressive induction with prolonged light exposure, whereas no significant changes were observed in light-exposed WT plants. The transgenic plants also expressed higher levels of most carotenoid biosynthesis pathway genes, including phytoene synthase (AtPSY), phytoene desaturase (AtPDS), lycopene-β-cyclase (AtLYCB), and ζ-carotene desaturase (AtZDS). The elongated hypocotyl 5 (HY5) and succinate dehydrogenase (SDH) genes were significantly induced following exposure to high light conditions for 12h, whereas phytochrome-interacting factor 7 (PIF7) was significantly downregulated in these plants.

Quantitative trait locus mapping analysis of multiple traits when using genotype data with potential errors.

BackgroundQuantitative trait locus (QTL) analysis aims to locate and estimate the effects of the genes influencing quantitative traits and infer the relationship between gene variants and changes in phenotypic characteristics using statistical methods. Some methods have been developed to map QTLs of multiple traits in the case of no genotype error in a given dataset. However, practical genetic data that people use may contain some potential errors because of the limitations of biotechnology. Common genetic data correction methods can only reduce errors, but cannot calculate the degree of error. In this paper, we propose a QTL mapping strategy for multiple traits in the presence of genotype errors.MethodsThe additive effect, dominant effect, recombination rate, error rate, and other parameters of QTLs can be simultaneously obtained using this new method in the framework of multiple-interval mapping.ResultsOur simulation results show that the accuracy of parameter estimation can be improved by considering the errors of marker genotypes during the analysis of genetic data. Real data analysis also shows that the new method proposed in this paper can map the QTLs of multiple traits more accurately.

Automated Imaging, Tracking, and Analytics Pipeline for Differentiating Environmental Effects on Root Meristematic Cell Division.

Exposure of plants to abiotic stresses, whether individually or in combination, triggers dynamic changes to gene regulation. These responses induce distinct changes in phenotypic characteristics, enabling the plant to adapt to changing environments. For example, iron deficiency and heat stress have been shown to alter root development by reducing primary root growth and reducing cell proliferation, respectively. Currently, identifying the dynamic temporal coordination of genetic responses to combined abiotic stresses remains a bottleneck. This is, in part, due to an inability to isolate specific intervals in developmental time where differential activity in plant stress responses plays a critical role. Here, we observed that iron deficiency, in combination with temporary heat stress, suppresses the expression of iron deficiency-responsive pPYE::LUC (POPEYE::luciferase) and pBTS::LUC (BRUTUS::luciferase) reporter genes. Moreover, root growth was suppressed less under combined iron deficiency and heat stress than under either single stress condition. To further explore the interaction between pathways, we also created a computer vision pipeline to extract, analyze, and compare high-dimensional dynamic spatial and temporal cellular data in response to heat and iron deficiency stress conditions at high temporal resolution. Specifically, we used fluorescence light sheet microscopy to image Arabidopsis thaliana roots expressing CYCB1;1:GFP, a marker for cell entry into mitosis, every 20 min for 24 h exposed to either iron sufficiency, iron deficiency, heat stress, or combined iron deficiency and heat stress. Our pipeline extracted spatiotemporal metrics from these time-course data. These metrics showed that the persistency and timing of CYCB1;1:GFP signal were uniquely different under combined iron deficiency and heat stress conditions versus the single stress conditions. These metrics also indicated that the spatiotemporal characteristics of the CYCB1;1:GFP signal under combined stress were more dissimilar to the control response than the response seen under iron deficiency for the majority of the 24-h experiment. Moreover, the combined stress response was less dissimilar to the control than the response seen under heat stress. This indicated that pathways activated when the plant is exposed to both iron deficiency and heat stress affected CYCB1;1:GFP spatiotemporal function antagonistically

Biological relevance of antler, horn, and pronghorn size in records programs

AbstractLong-term datasets are becoming increasingly important for assessing population- and species-level responses to a changing environment. Programs that record morphological measurements of horns, antlers, and pronghorns were established in the early- to mid-20th century to collect biological information about animals that possess large horns, antlers, or pronghorns, which could be used to assess the effectiveness of conservation efforts for large mammals in North America. The general relevance of record books has been questioned because of the minimum size requirements for inclusion in a record book, which may mask trends when changes in the population occur. We compared trends in size of antlers, horns, and pronghorns through time using records from three records programs with different minimum size requirements to evaluate the influence of entry requirements on temporal trends. We also investigated whether horn, antler, or pronghorn size affected the probability of specimens being submitted to a records program. Only two of 17 categories exhibited less-pronounced trends in the record book with the highest size requirements for entry, and in two categories trends were more pronounced. Although societal interest in submitting eligible specimens increased slightly over time in one of six categories, the probability of voluntary entry was largely random and not affected by year of harvest or size of specimen. In contrast to previous criticisms, trends in record books should not be expected to represent the size of all males within a population. Instead, our evaluation indicates that the records programs we examined can provide a useful resource for assessing long-term changes in phenotypic characteristics of ungulates, but importantly, they represent the respective range of sizes within which each program collects data.

EFFECTS OF EXOGENOUS LACTOFERRIN ON PHENOTYPIC PROFILE AND INVASIVENESS OF HUMAN PROSTATE CANCER CELLS (DU-145 AND LNCaP) IN VITRO

To investigate the biological effects of exogenous lactoferrin (LF) on phenotypic profile and invasiveness of human prostate cancer(PC) cells in vitro. Human PC cell lines (LNCaP, DU-145) were cultured with an exogenous LF at a dose corresponding to IC30. The expression levels of steroid hormone receptors (androgen receptor, estrogen receptor, progesterone receptor), Her2/neu, Ki-67, E- and N-cadherin, were monitored by immunohistochemical analysis. The levels of miRNAs were assessed using q-PCR. The invasive activity of the cells was examined in a standard invasion test. Exogenous LF reduced expression of steroid hormone receptors (ERα and PR) and Ki-67in both PC cell lines. The expression of E-cadherin increased significantly in LF-treated DU-145cells. Also, we established the decrease in invasive activity upon LF treatment by 40% and 30% in DU-145and LNCaP cells, respectively. In DU-145cells, incubation with exogenous LF resulted in an increase in the expression of oncosuppressive (miR-133a and miR-200b) miRNAs. Exogenous LF causes the changes in phenotypic characteristics of PC cells and levels of oncogenic and oncosuppressive miRNAs involved in the regulation of key cellular processes.

Reduction in DNA topoisomerase I level affects growth, phenotype and nucleoid architecture of Mycobacterium smegmatis.

The steady-state negative supercoiling of eubacterial genomes is maintained by the action of DNA topoisomerases. Topoisomerase distribution varies in different species of mycobacteria. While Mycobacterium tuberculosis (Mtb) contains a single type I (TopoI) and a single type II (Gyrase) enzyme, Mycobacterium smegmatis (Msm) and other members harbour additional relaxases. TopoI is essential for Mtb survival. However, the necessity of TopoI or other relaxases in Msm has not been investigated. To recognize the importance of TopoI for growth, physiology and gene expression of Msm, we have developed a conditional knock-down strain of TopoI in Msm. The TopoI-depleted strain exhibited extremely slow growth and drastic changes in phenotypic characteristics. The cessation of growth indicates the essential requirement of the enzyme for the organism in spite of having additional DNA relaxation enzymes in the cell. Notably, the imbalance in TopoI level led to the altered expression of topology modulatory proteins, resulting in a diffused nucleoid architecture. Proteomic and transcript analysis of the mutant indicated reduced expression of the genes involved in central metabolic pathways and core DNA transaction processes. RNA polymerase (RNAP) distribution on the transcription units was affected in the TopoI-depleted cells, suggesting global alteration in transcription. The study thus highlights the essential requirement of TopoI in the maintenance of cellular phenotype, growth characteristics and gene expression in mycobacteria. A decrease in TopoI level led to altered RNAP occupancy and impaired transcription elongation, causing severe downstream effects.

Pathological vitreous causes cell line‐derived (but not donor‐derived) retinal pigment epithelial cells to display proliferative vitreoretinopathy‐like features in culture

It is well understood that epithelial mesenchymal transformation occurs when retinal pigment epithelial cells, sourced from either a cell line or cadaver eye, are cultured in the presence of cadaver-derived vitreous. We sought to study the changes in retinal pigment epithelial cells when cell line-derived retinal pigment epithelial cells are cultured in the presence of pathological vitreous. Prospective study. 42 patients with rhegmatogenous retinal detachments. D407 retinal pigment epithelial cells were cultured in the presence of cadaver-derived vitreous or vitreous/subretinal fluid derived from patients undergoing retinal reattachment surgeries. Besides the changes in phenotypic characteristics, the viability, proliferation, migration, mesenchymal marker expression and changes in the extracellular matrix components were also evaluated. Fibrotic phenotype in cell culture. Our study clearly demonstrates that cell line-derived retinal pigment epithelial cells (unlike donor-derived retinal pigment epithelial cells) cultured in the presence of patient-derived vitreous/subretinal fluid, exhibit characteristic features of proliferative vitreoretinopathy. We propose that it is the synergistic effect of the combined use of (i) pathological vitreous, rather than cadaver-derived vitreous (since rhegmatogenous retinal detachment-derived pathological vitreous and subretinal fluid contain exaggerated amounts of growth factors, which could predispose to proliferative vitreoretinopathy development) and (ii) cells from an immortal cell culture (cell line), rather than from primary cell cultures (since cells subjected to continuous serial passaging acquire some mesenchymal characteristics), which together result in not only a unique phenotype, but also prime these cells towards display of features associated with proliferative vitreoretinopathy.

Estabilidade fenotípica de genótipos de morangueiro submetidos a número variável de subcultivos in vitro

Objetivando analisar o número de subcultivos que permita a multiplicação do morangueiro, sem que ocorram alterações nas características fenotípicas dos clones submetidos a esse processo, explantes dos genótipos 'Aromas', 'Camarosa' e 'Camino Real' foram submetidos, em uma primeira etapa, a 12 ciclos de subcultivos in vitro e, no ano seguinte, explantes dos mesmos genótipos, das mesmas plantas matrizes, foram subcultivados por três ciclos. Após a fase de aclimatização, as mudas foram transplantadas para canteiros, e o experimento conduzido segundo delineamento em blocos casualizados em esquema fatorial 3x2 (genótipos e níveis de subcultivos). As características avaliadas foram: altura da planta e comprimento da raiz (cm), massas fresca e seca da parte aérea e da raiz (g) e número de estolões por planta. Na avaliação a campo das características fenotípicas, não foram observadas diferenças significativas entre os dois níveis de subcultivos. As cultivares 'Camarosa' e 'Camino Real' apresentaram maior variação entre os dois níveis de subcultivos, sendo as maiores variações observadas em plantas submetidas a três subcultivos. Com 12 subcultivos in vitro de plantas de morangueiro, das cultivares 'Aromas', 'Camarosa' e 'Camino Real', é possível obter maior número de mudas micropropagadas sem alterações nas características fenotípicas. A cultivar 'Aromas' apresentou uma maior estabilidade nas características fenotípicas estudadas em relação aos diferentes ciclos de subcultivo in vitro.

Characterizing Vancomycin-Resistant Enterococcus Strains with Various Mechanisms of Daptomycin Resistance Developed in an In Vitro Pharmacokinetic/Pharmacodynamic Model

Two daptomycin (DAP) regimens were evaluated in a pharmacokinetic/pharmacodynamic (PK/PD) model, and the mutants recovered were examined for changes in phenotypic characteristics. Three Enterococcus faecium strains (vancomycin-resistant Enterococcus [VRE] ATCC 51559, VRE 12311, and VRE SF 12047) were utilized in a 7-day, 1-compartment in vitro PK/PD model. The simulated dosing regimens were DAP at 6 mg/kg/day (free C(max) [fC(max)] = 7.9 μg/ml, half-life [t(1/2)] = 8 h) and DAP at 10 mg/kg/day (fC(max) = 13.17 μg/ml, t(1/2) = 8 h). Samples were plated daily on Mueller-Hinton agar containing DAP at 16 μg/ml and 50 mg/liter Ca(2+) to assess the emergence of DAP resistance. For each strain, the mutant with the highest DAP MIC was then evaluated for changes in relative surface charge, cell wall thickness, and cytoplasmic membrane depolarization induced by DAP. The initial DAP MICs were 4 μg/ml for all 3 strains. A dose-dependent response and regrowth were observed for DAP 6 mg/kg/day and DAP 10 mg/kg/day against all 3 strains. Mutants of VRE ATCC 51559 (MIC = 128 and 64 μg/ml) and VRE 12311 (MIC = 256 and 32 μg/ml) were recovered from the DAP 6 mg and DAP 10 mg regimen, respectively. For VRE SF 12047, a mutant (MIC = 64 μg/ml) was recovered from the DAP 6 mg model. All mutants displayed an increase in relative surface charge compared to those of their respective parent strains. The DAP-resistant mutants displayed a 43 to 58% increase in cell wall thickness (P < 0.0001), while DAP membrane depolarization decreased by 53 to 65% compared to that of the susceptible strains. VRE with DAP resistance displayed increased surface charge, increased cell wall thickness, and decreased depolarization induced by DAP, consistent with previous observations in Staphylococcus aureus with reduced DAP susceptibility. Further characterization of DAP-resistant VRE is warranted.

Gene-associated Single Nucleotide Polymorphism (SNP) in Cinnamate 4-Hydroxylase (C4H) and Cinnamyl Alcohol Dehydrogenase (CAD) Genes from Acacia mangium Superbulk Trees

Candidate-gene-based association study which involves the identification of causative Single Nucleotide Polymorphisms (SNPs) for excellent traits has been proposed as a promising approach to dissect complex traits in forest trees. Hence, the goal of this study was to identify the genetic association among SNPs from Cinnamate 4-Hydroxylase (C4H) and Cinnamyl Alcohol Dehydrogenase (CAD) genes and an array of wood properties namely, specific gravity, wood density, fiber-length, cell wall thickness and microfibril angle from Acacia mangium Superbulk trees. Sequence variations within these two genes in 12 A. mangium Superbulk trees were examined and wood properties were measured. The data obtained was tested using General Linear Model (GLM) within TASSEL software. Two SNPs were identified in the exon of C4H, of which all the SNPs caused nonsynonymous mutations whereas five SNPs were identified in the CAD exons along with one deletion mutation. In addition, two SNPs were also identified in the CAD introns. Variation in these two lignin biosynthesis genes might change the structural, functional or biochemical properties of the enzyme being produced, and therefore possibly lead to changes in phenotypic characteristic of the trees. The genetic association study also revealed that SNPs in CAD gene do associate with the wood density, specific gravity and cell wall thickness (p<0.05). However, no significant results were obtained for SNPs in C4H gene with wood properties studied. Thickening of cell wall is affected by the arrangement of biopolymer aggregates which comprise of cellulose, hemicellulose and lignin. Results indicated that SNP in CAD gene might alter the lignin biosynthesis and thus lead to changes in phenotypic characteristics of the trees. Overall, the study has demonstrated that SNP is very useful in association genetic study to identify Quantitative Trait Nucleotide (QTN) which then leads to Gene-assisted Selection (GAS) in the tree breeding programme.

Divergent Squamous Differentiation in Upper Urothelial Carcinoma—Comparative Clinicopathological and Molecular Study

Upper urothelial carcinoma (UUC) has a plasticity to demonstrate divergent differentiation with squamous metaplastic elements. There was no previous study exploring profiling of molecular markers in metaplastic squamous upper urothelial carcinoma (SUUC) and conventional upper urothelial carcinoma (CUUC). The aims of this study was to compare expression of the phenotypic characteristics of tumors and molecular markers (p53, p16, cyclin D1, E-cadherin, HER-2, Ki-67, Bcl-2, Bax) in SUUC and CUUC. SUUC was detected in 20% of 44 patients. There was significant difference between SUUC and CUUC in the pathological stage, grade, growth and presence of lymphovasular invasion (p < 0.05; 0.05; 0.05; 0.01 respectively). The mean Ki-67 and p53 labeling index was significantly higher in SUUC than in CUUC (p < 0.05; 0.05). There was no significant difference in the expression of p16, cyclin D1, E-cadherin, HER-2, Bcl-2 and Bax between SUUC and CUUC. Univariant model showed that SUUC was significantly associated with lymphovascular invasion (p = 0.007), Ki-67 activity (p = 0.016) and growth (p = 0.026). Exploration of UUC with squamous divergent differentiation showed changes in phenotypic characteristics and Ki-67, as well as similar molecular profile with CUUC.

Effect of broad-spectrum matrix metalloproteinase inhibition on atherosclerotic plaque stability

Matrix metalloproteinases (MMPs) form a large family of enzymes that collectively can degrade all components of the extracellular matrix, and there is widespread interest in developing MMP inhibitors for the prevention of atherosclerotic plaque rupture. We have therefore investigated the effects of a broad-spectrum MMP inhibitor, RS-130830, on plaque development and stability. This compound inhibits a wide range of MMPs at concentrations below 20 nmol/L. Apolipoprotein E knockout mice were fed a Western diet. Dietary administration of RS-130830 commenced at the same time as fat-feeding and continued for 8, 12, 26 or 36 weeks. To investigate the effect of RS-130830 on established plaques, mice were fed high-fat diet for 16 weeks before initiation of drug treatment and were terminated 20 weeks after this. Broad-spectrum MMP inhibition was associated with a significant increase in plaque area, but there was no change in the incidence of plaque rupture. There were unfavourable changes in phenotypic characteristics associated with plaque instability, such as an increased lipid content and decreased collagen content. These data suggest that broad-spectrum MMP inhibition RS-130830 does not have a beneficial effect on atherosclerosis in the apolipoprotein E knockout mouse model, and indicate that more selective compounds would be preferable.

A 10-year follow-up study of penicillin-non-susceptible S. pneumoniae during an intervention programme in Malmö, Sweden

Changes in the proportion of penicillin-non-susceptible Streptococcus pneumoniae (PNSP) isolates during an intervention programme were evaluated by phenotypic analysis of all initial isolates with penicillin MIC > or =0.5 microg/ml (n=1248) collected 1995-2004. During the study period, the proportion of such isolates was fairly constant (12-19%), and there was no statistically significant variation in the proportion of total PNSP cases (MIC > or =0.12 microg/ml) or PNSP with MIC > or =0.5 microg/ml, with the exception of an increase in 2004. Analysis restricted to clinical cases revealed no statistically significant changes. 23 different serogroups were found, and serogroup 9 isolates accounted for almost half of the PNSP cases. Only minor changes in phenotypic characteristics occurred in the other serogroups, which indicates that the increase in PNSP in 2004 was not due to import of a new resistant clone. Antibiotic consumption is considered to be an important risk factor for penicillin resistance in S. pneumoniae. After initiation of the intervention programme in Malmö, overall prescribing of antibiotics decreased 28%, and the reduction was even greater among children (52%). In conclusion, the proportion of PNSP isolates in Malmö has remained stable, despite the intervention programme and decreased consumption of antibiotics.

Molecular Characterization And Comparison Of Multi Drug Resistant Strains Of Mycobacterium Tuberculosis By Phenotypic And Genotypic Method

Introduction: Drug resistant tuberculosis is a signifi cant threat to tuberculosis control because only a few effective drugs are available against M. tuberculosis. The aims of this study were to compare multi drug resistant (MDR) strains of tuberculosis by phenotypic and genotypic method and determine type, location and frequency of rpoB and KatG gene mutations. Methodology: Anti-tuberculosis drugs susceptibility test of M. tuberculosis grown on Lowenstein Jensen medium was performed by proportion method. MDR cases were analyzed for mutation of rpoB and KatG genes. The regions of these genes were amplifi ed by polymerase chain reaction (PCR) and sequenced. Results: Two different mutations were identifi ed in rifampicin resistant strains. The most common point mutations were in codons TCG 531→TTG (85%) and GAC 516→TTC (15%) of the rpoB gene. Two different mutations in KatG gene were detected. The most common KatG point mutations were AGC 315 ACC (Ser→Thr) (85%) and CGG 463 CTG (Arg→Leu) (10%). In this study DNA sequencing analysis did not fi nd mutation on KatG gene of one of the strain tested. Male and female were equally affected by MDR tuberculosis and majorities (35%) of them were found in 21-30 years age group. Conclusion: The present investigation agrees that genetic mutation is responsible for change in phenotypic characteristics of M. Tuberculosis. DOI: http://dx.doi.org/10.3126/saarctb.v8i2.5896 SAARCTB 2011; 8(2): 17-22