Expression of the Arabidopsis thaliana BBX32 Gene in Soybean Increases Grain Yield

Crop yield is a highly complex quantitative trait. Historically, successful breeding for improved grain yield has led to crop plants with improved source capacity, altered plant architecture, and increased resistance to abiotic and biotic stresses. To date, transgenic approaches towards improving crop grain yield have primarily focused on protecting plants from herbicide, insects, or disease. In contrast, we have focused on identifying genes that, when expressed in soybean, improve the intrinsic ability of the plant to yield more. Through the large scale screening of candidate genes in transgenic soybean, we identified an Arabidopsis thaliana B-box domain gene (AtBBX32) that significantly increases soybean grain yield year after year in multiple transgenic events in multi-location field trials. In order to understand the underlying physiological changes that are associated with increased yield in transgenic soybean, we examined phenotypic differences in two AtBBX32-expressing lines and found increases in plant height and node, flower, pod, and seed number. We propose that these phenotypic changes are likely the result of changes in the timing of reproductive development in transgenic soybean that lead to the increased duration of the pod and seed development period. Consistent with the role of BBX32 in A. thaliana in regulating light signaling, we show that the constitutive expression of AtBBX32 in soybean alters the abundance of a subset of gene transcripts in the early morning hours. In particular, AtBBX32 alters transcript levels of the soybean clock genes GmTOC1 and LHY-CCA1-like2 (GmLCL2). We propose that through the expression of AtBBX32 and modulation of the abundance of circadian clock genes during the transition from dark to light, the timing of critical phases of reproductive development are altered. These findings demonstrate a specific role for AtBBX32 in modulating soybean development, and demonstrate the validity of expressing single genes in crops to deliver increased agricultural productivity

Ultrasound Biomicroscopy for Longitudinal Studies of Carotid Plaque Development in Mice: Validation with Histological Endpoints

Atherosclerosis is responsible for the death of thousands of Americans each year. The carotid constriction model of plaque development has recently been presented as a model for unstable plaque formation in mice. In this study we 1) validate ultrasound biomicroscopy (UBM) for the determination of carotid plaque size, percent stenosis, and plaque development in live animals, 2) determine the sensitivity of UBM in detecting changes in blood flow induced by carotid constriction and 3) test whether plaque formation can be predicted from blood flow parameters measured by UBM. Carotid plaques were induced by surgical constriction in Apo E-/- mice. Arteries were imaged bi-weekly by UBM, at which time PW-Doppler measurements of proximal blood flow, as well as plaque length and percent stenosis were determined. Histology was performed 9 weeks post surgery. When compared to whole mount post-mortem measurements, UBM accurately reported carotid plaque length. Percent stenosis, based on transverse B-mode UBM measurements, correlated well with that calculated from histological sections. PW-Doppler revealed that constriction reduced maximum systolic velocity (v(max)) and duration of the systolic velocity peak (t(s)/t(t)). Pre-plaque (2 week post-surgery) PW-Doppler parameters (v(max) and t(s)/t(t)) were correlated with plaque length at 9 weeks, and were predictive of plaque formation. Correlation of initiating PW-Doppler parameters (v(max) and t(s)/t(t)) with resulting plaque length established the degree of flow disturbance required for subsequent plaque development and demonstrated its power for predicting plaque developmen

The effect of engineered nanotopography of electrospun microfibers on fiber rigidity and macrophage cytokine production

<p>Currently, it is unknown how the mechanical properties of electrospun fibers, and the presentation of surface nanotopography influence macrophage gene expression and protein production. By further elucidating how specific fiber properties (mechanical properties or surface properties) alter macrophage behavior, it may be possible to create electrospun fiber scaffolds capable of initiating unique cellular and tissue responses. In this study, we determined the elastic modulus and rigidity of fibers with varying topographies created by finely controlling humidity and including a non-solvent during electrospinning. In total,five fiber scaffold types were produced. Analysis of fiber physical properties demonstrated no change in fiber diameter amongst the five different fiber groups. However, the four different fibrous scaffolds with nanopits or divots each possessed different numbers of pits with different nanoscale dimensions. Unpolarized bone marrow derived murine macrophages (M0), macrophages polarized towards a pro-inflammatory phenotype (M1), or macrophages polarized towards anti-inflammatory phenotype (M2b) were placed onto each of the scaffolds and cytokine RNA expression and protein production were analyzed. Specific nanotopographies did not appreciably alter cytokine production from undifferentiated macrophages (M0) or anti-inflammatory macrophages (M2b), but a specific fiber (with many small pits) did increase IL-12 transcript and IL-12 protein production compared to fibers with small divots. When analyzing the mechanical properties between fibers with divots or with many small pits,divoted fibers possessed similar elastic moduli but different stiffness values. In total,we present techniques capable of creating unique electrospun fibers. These unique fibers have varying fiber mechanical characteristics and modestly modulate macrophage cytokine expression.</p

Percent stenosis calculated from UBM parameters correlates with histological measurements.

<p>(a) Transverse B-mode image of RCCA 2 and 9 weeks after surgery. Upper Images, Adventitia is marked “A”, plaque “P”, “Medial layer M”. Lower, images are identical, but the inner elastic lamina is outlined with dotted white line, lumen with a red dotted line. Trichrome-stained transverse sections of the same 9 wk artery; internal elastic lamina (IEL) and lumen are outlined in white and red, respectively. (b) IMT measurements of casted and cuffed animals after 2 and 9 weeks of surgery. n.s. Not significant. (c) Linear regression analysis of percent stenosis estimated by UBM and histology. See Materials and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0029944#s2" target="_blank">Methods</a> for calculations of percent stenosis. Dotted lines indicate 95% confidence intervals; correlation coefficient = 0.75, n = 10 casts. (d) Comparison of UBM and histology measurements using Bland-Altman analysis. Dotted lines indicate 95% limits of agreement. Bias is indicated by solid line, was calculated as −4.76 mm±13.3 SD.</p

Plaques Progress More Rapidly in Casted vs Cuffed Animals.

<p>The rate of plaque growth was determined by performing individual linear regression analysis of plaque length or stenosis over time for each animal. The slopes (rates) were compared by unpaired t-test. (a) Rate of plaque growth in length (b) Rate of plaque growth in terms of stenosis. Data are mean±SEM, n = 7 cuffs, 10 casts “*” indicates p<0.05. (See <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0029944#pone.0029944.s002" target="_blank">Fig. S2</a> for source data.)</p

Changes in Lumen and IEL area upon carotid constriction.

<p>Data was analyzed by One Way ANOVA.</p><p>“*”indicates p<0.05 as compared to presurgery.</p>“†”<p>indicates p<0.05 as compared to 2 weeks post surgery.</p

Sensitivity and Specificity of PW-Doppler parameters t_s/t_T and v_max as predictors of plaque formation.

<p>Sensitivity and Specificity of PW-Doppler parameters t_s/t_T and v_max as predictors of plaque formation.</p