Stomatal movements, which regulate gas exchange in plants, involve pronounced changes in the shape and volume of the guard cell. To test whether the changes are regulated by actin filaments, we visualized microfilaments in mature guard cells and examined the effects of actin antagonists on stomatal movements. Immunolocalization on fixed cells and microinjection of fluorescein isothiocyanate-phalloidin into living guard cells of Commelina communis L. showed that cortical microfilaments were radially distributed, fanning out from the stomatal pore site, resembling the known pattern of microtubules. Treatment of epidermal peels with phalloidin prior to stabilizing microfilaments with m-maleimidobenzoyl N-hydroxysuccimimide caused dense packing of radial microfilaments and an accumulation of actin around many organelles. 60th stomatal closing induced by abscisic acid and opening under light were inhibited. Treatment of guard cells with cytochalasin D abolished the radial pattern of microfilaments; generated sparse, poorly oriented arrays; and caused partia1 opening of dark-closed stomata. These results suggest that microfilaments participate in stomatal aperture regulation.

Biocatalytic cyclization is highly desirable for efficient synthesis of biologically derived chemical substances, such as the commodity chemicals ε-caprolactam and δ-valerolactam. To identify biocatalysts in lactam biosynthesis, we develop a caprolactam-detecting genetic enzyme screening system (CL-GESS). The Alcaligenes faecalis regulatory protein NitR is adopted for the highly specific detection of lactam compounds against lactam biosynthetic intermediates. We further systematically optimize the genetic components of the CL-GESS to enhance sensitivity, achieving 10-fold improvement. Using this highly sensitive GESS, we screen marine metagenomes and find an enzyme that cyclizes ω-amino fatty acids to lactam. Moreover, we determine the X-ray crystal structure and catalytic residues based on mutational analysis of the cyclase. The cyclase is also used as a helper enzyme to sense intracellular ω-amino fatty acids. We expect this simple and accurate biosensor to have wide-ranging applications in rapid screening of new lactam-synthesizing enzymes and metabolic engineering for lactam bio-production.

1, †DICE1 (deleted in cancer 1), first identified in human lung carcinoma cell lines, is a candidate tumor suppressor, but the details of its activity remain largely unknown. We have found that RNA interference of its C. elegans homolog (DIC-1) produced inviable embryos with increased apoptosis, cavities in cells and abnormal morphogenesis. In the dic-1(RNAi) germ line, ced-3-dependent apoptosis increased, and cell cavities appeared at the late-pachytene/oogenic stage, leading to defective oogenesis. Immunofluorescence microscopy of DIC-1 revealed its ubiquitous expression in the form of cytoplasmic foci, and cryoelectron microscopy narrowed down the location of the foci to the inner membrane of mitochondria. After dic-1 RNAi, mitochondria had an irregular morphology and contained numerous internal vesicles. Homozygous embryos from a heterozygous dic-1 mother arrested at the L3 larval stage, in agreement with the essential role of DIC-1 in mitochondria. In summary, C. elegans DIC-1 plays a crucial role in the formation of normal morphology of the mitochondrial cristae/inner membrane. Our results suggest that human DICE1 may have several functions in multiple intracellular locations.

Pressure drop (p) and collection efficiency (η) are used to evaluate the separation performance of the cyclone separator. In this study, we conducted comparative study of cyclone models using response surface methodology (RSM), back propagation neural network (BPNN), and group method of data handling (GMDH) networks to develop optimal predictive cyclone models. Also, we conducted multi-objective optimization for maximizing model and minimizing model using genetic algorithm (GA). CFD was performed instead of experimental method to get the estimated values for modeling of p and η. The validation results of CFD showed 0.5% and 2% errors for p and η, respectively, compared with the experimental data. Second, design of experiment (DOE) analysis for 10 cyclone geometrical parameters was executed to obtain the significant geometrical parameters. Vortex finder diameter D x , inlet width a, inlet height b and cone height H co have a significant effect on η and p. However, interaction effects between the geometrical parameters have small effects. The cyclone models by RSM, BPNN and GMDH based on 25 CFD training set were developed. The predictive performance results by the cyclone models were compared by 25 CFD test set. The GMDH method achieved the best prediction for p (R 2 = 99.7, RMSE = 0.102) R 2 adjusted = 98.99, RMSE = 0.0119) than the RSM, BPNN cyclone models. Additionally, uncertainty analysis was performed to estimate the quantitative performance of cyclone models. The results show that the uncertainty width of GMDH models achieved the best prediction (η: ±0.0065, p: ±0.0188). Finally, GA was applied to optimize the GMDH models simultaneously. GA generated 70 non-dominant solutions. Reproducibility of five optimal points was validated by using CFD. The trade-off optimal point showed improvement by 24.31%, 18% and 8.79% for p d 50 and η, respectively.

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