A Quantitative and Dynamic Model of the Arabidopsis Flowering Time Gene Regulatory Network

Various environmental signals integrate into a network of floral regulatory genes leading to the final decision on when to flower. Although a wealth of qualitative knowledge is available on how flowering time genes regulate each other, only a few studies incorporated this knowledge into predictive models. Such models are invaluable as they enable to investigate how various types of inputs are combined to give a quantitative readout. To investigate the effect of gene expression disturbances on flowering time, we developed a dynamic model for the regulation of flowering time in Arabidopsis thaliana. Model parameters were estimated based on expression time-courses for relevant genes, and a consistent set of flowering times for plants of various genetic backgrounds. Validation was performed by predicting changes in expression level in mutant backgrounds and comparing these predictions with independent expression data, and by comparison of predicted and experimental flowering times for several double mutants. Remarkably, the model predicts that a disturbance in a particular gene has not necessarily the largest impact on directly connected genes. For example, the model predicts that SUPPRESSOR OF OVEREXPRESSION OF CONSTANS (SOC1) mutation has a larger impact on APETALA1 (AP1), which is not directly regulated by SOC1, compared to its effect on LEAFY (LFY) which is under direct control of SOC1. This was confirmed by expression data. Another model prediction involves the importance of cooperativity in the regulation of APETALA1 (AP1) by LFY, a prediction supported by experimental evidence. Concluding, our model for flowering time gene regulation enables to address how different quantitative inputs are combined into one quantitative output, flowering time.     

4-(2-pyridyl)piperazine-1-carboxamides: potent vanilloid receptor 1 antagonists

A series of 4-(2-pyridyl)piperazine-1-carboxamide analogues based on the lead compound 1 was synthesized and evaluated for VR1 antagonist activity in capsaicin-induced (CAP) and pH (5.5)-induced (pH) FLIPR assays in a rat VR1-expressing HEK293 cell line. Potent VR1 antagonists were identified through SAR studies. From these studies, 18 was found to be very potent in the in vitro assay [IC(50)=4.8 nM (pH) and 35 nM (CAP)] and orally available in rat (F%=15.1).     

Population size and identity influence the reaction norm of the rare,endemic plant Cochlearia bavarica across a gradient of environmental stress

Habitat degradation and loss can result in population decline and genetic erosion, limiting the ability of organisms to cope with environmental change, whether this is through evolutionary genetic response (requiring genetic variation) or through phenotypic plasticity (i.e., the ability of a given genotype to express a variable phenotype across environments). Here we address the question whether plants from small populations are less plastic or more susceptible to environmental stress than plants from large populations. We collected seed families from small (<100) versus large natural populations (>1,000 flowering plants) of the rare, endemic plant Cochlearia bavarica (Brassicaceae). We exposed the seedlings to a range of environments, created by manipulating water supply and light intensity in a 2 x 2 factorial design in the greenhouse. We monitored plant growth and survival for 300 days. Significant effects of offspring environment on offspring characters demonstrated that there is phenotypic plasticity in the responses to environmental stress in this species. Significant effects of population size group, but mainly of population identity within the population size groups, and of maternal plant identity within populations indicated variation due to genetic (plus potentially maternal) variation for offspring traits. The environment x maternal plant identity interaction was rarely significant, providing little evidence for genetically- (plus potentially maternally-) based variation in plasticity within populations. However, significant environment x population-size-group and environment x population-identity interactions suggested that populations differed in the amount of plasticity, the mean amount being smaller in small populations than in large populations. Whereas on day 210 the differences between small and large populations were largest in the environment in which plants grew biggest (i.e., under benign conditions), on day 270 the difference was largest in stressful environments. These results show that population size and population identity can affect growth and survival differently across environmental stress gradients. Moreover, these effects can themselves be modified by time-dependent variation in the interaction between plants and their environment.     

Stealth proteins: in silico identification of a novel protein family rendering bacterial pathogens invisible to host immune defense

There are a variety of bacterial defense strategies to survive in a hostile environment. Generation of extracellular polysaccharides has proved to be a simple but effective strategy against the host's innate immune system. A comparative genomics approach led us to identify a new protein family termed Stealth, most likely involved in the synthesis of extracellular polysaccharides. This protein family is characterized by a series of domains conserved across phylogeny from bacteria to eukaryotes. In bacteria, Stealth (previously characterized as SacB, XcbA, or WefC) is encoded by subsets of strains mainly colonizing multicellular organisms, with evidence for a protective effect against the host innate immune defense. More specifically, integrating all the available information about Stealth proteins in bacteria, we propose that Stealth is a D-hexose-1-phosphoryl transferase involved in the synthesis of polysaccharides. In the animal kingdom, Stealth is strongly conserved across evolution from social amoebas to simple and complex multicellular organisms, such as Dictyostelium discoideum, hydra, and human. Based on the occurrence of Stealth in most Eukaryotes and a subset of Prokaryotes together with its potential role in extracellular polysaccharide synthesis, we propose that metazoan Stealth functions to regulate the innate immune system. Moreover, there is good reason to speculate that the acquisition and spread of Stealth could be responsible for future epidemic outbreaks of infectious diseases caused by a large variety of eubacterial pathogens. Our in silico identification of a homologous protein in the human host will help to elucidate the causes of Stealth-dependent virulence. At a more basic level, the characterization of the molecular and cellular function of Stealth proteins may shed light on fundamental mechanisms of innate immune defense against microbial invasion.     

Residue size at position 87 of cytochrome P450 BM-3 determines its stereoselectivity in propylbenzene and 3-chlorostyrene oxidation.

We report here oxidation of propylbenzene and 3-chlorostyrene by wild-type cytochrome P450 BM-3 with high turnover (479 nmol 1-phenyl-1-propanol/min/nmol P450 and 300 nmol 3-chlorostyrene oxide/min/nmol P450). Furthermore, the residue size at position 87 of P450 BM-3 was found to play critical roles in determining stereoselectivity in oxidation of propylbenzene and 3-chlorostyrene. Replacement of Phe87 with Val, Ala and Gly resulted in decreases in optical purity of produced (R)-(+)-1-phenyl-1-propanol from 90.0 to 37.4, 26.0 and -15.6% e.e., respectively, and in increases in those of produced (R)-(+)-3-chlorostyrene oxide from -61.0 to -38.0, 67.0 and 94.6% e.e., respectively.     

Effect of Different Saccharides on Viability of Isolated Microspores and Androgenic Induction in Zea Mays

Sucrose, glucose, fructose, and melibiose in different concentrations and combinations in the induction media influenced the viability of the isolated maize microspores and the formation of multinuclear structures. The induction of multinuclear structures on media containing combination of sucrose, fructose and glucose was lower than on media only with sucrose. In media containing melibiose alone or in combination with sucrose, no induction of multinuclear structures was found, however, microspore viability was improved.     

Equivalent genetic roles for bmp7/snailhouse and bmp2b/swirl in dorsoventral pattern formation

A bone morphogenetic protein (BMP) signaling pathway acts in the establishment of the dorsoventral axis of the vertebrate embryo. Here we demonstrate the genetic requirement for two different Bmp ligand subclass genes for dorsoventral pattern formation of the zebrafish embryo. From the relative efficiencies observed in Bmp ligand rescue experiments, conserved chromosomal synteny, and isolation of the zebrafish bmp7 gene, we determined that the strongly dorsalized snailhouse mutant phenotype is caused by a mutation in the bmp7 gene. We show that the original snailhouse allele is a hypomorphic mutation and we identify a snailhouse/bmp7 null mutant. We demonstrate that the snailhouse/bmp7 null mutant phenotype is identical to the presumptive null mutant phenotype of the strongest dorsalized zebrafish mutant swirl/bmp2b, revealing equivalent genetic roles for these two Bmp ligands. Double mutant snailhouse/bmp7; swirl/bmp2b embryos do not exhibit additional or stronger dorsalized phenotypes, indicating that these Bmp ligands do not function redundantly in early embryonic development. Furthermore, overexpression experiments reveal that Bmp2b and Bmp7 synergize in the ventralization of wild-type embryos through a cell-autonomous mechanism, suggesting that Bmp2b/Bmp7 heterodimers may act in vivo to specify ventral cell fates in the zebrafish embryo.