iCanPlot: visual exploration of high-throughput omics data using interactive Canvas plotting

Increasing use of high throughput genomic scale assays requires effective visualization and analysis techniques to facilitate data interpretation. Moreover, existing tools often require programming skills, which discourages bench scientists from examining their own data. We have created iCanPlot, a compelling platform for visual data exploration based on the latest technologies. Using the recently adopted HTML5 Canvas element, we have developed a highly interactive tool to visualize tabular data and identify interesting patterns in an intuitive fashion without the need of any specialized computing skills. A module for geneset overlap analysis has been implemented on the Google App Engine platform: when the user selects a region of interest in the plot, the genes in the region are analyzed on the fly. The visualization and analysis are amalgamated for a seamless experience. Further, users can easily upload their data for analysis--which also makes it simple to share the analysis with collaborators. We illustrate the power of iCanPlot by showing an example of how it can be used to interpret histone modifications in the context of gene expression.     

Cortactin as a Target for FAK in the Regulation of Focal Adhesion Dynamics

Background

Efficient cell movement requires the dynamic regulation of focal adhesion (FA) formation and turnover. FAs are integrin-associated sites of cell attachment and establish linkages to the cellular actin cytoskeleton. Cells without focal adhesion kinase (FAK), an integrin-activated tyrosine kinase, exhibit defects in FA turnover and cell motility. Cortactin is an actin binding adaptor protein that can influence FA dynamics. FAK and cortactin interact, but the cellular role of this complex remains unclear.

Principal Findings

Using FAK-null fibroblasts stably reconstituted with green fluorescent protein (GFP) tagged FAK constructs, we find that FAK activity and FAK C-terminal proline-rich region 2 (PRR2) and PRR3 are required for FA turnover and cell motility. Cortactin binds directly to FAK PRR2 and PRR3 sites via its SH3 domain and cortactin expression is important in promoting FA turnover and GFP-FAK release from FAs. FAK-cortactin binding is negatively-regulated by FAK activity and associated with cortactin tyrosine phosphorylation. FAK directly phosphorylates cortactin at Y421 and Y466 and over-expression of cortactin Y421, Y466, and Y482 mutated to phenylalanine (3YF) prevented FAK-enhanced FA turnover and cell motility. However, phospho-mimetic cortactin mutated to glutamic acid (3YE) did not affect FA dynamics and did not rescue FA turnover defects in cells with inhibited FAK activity or with PRR2-mutated FAK that does not bind cortactin.

Conclusions

Our results support a model whereby FAK-mediated FA remodeling may occur through the formation of a FAK-cortactin signaling complex. This involves a cycle of cortactin binding to FAK, cortactin tyrosine phosphorylation, and subsequent cortactin-FAK dissociation accompanied by FA turnover and cell movement.     

Identification of RAPD and SCAR markers associated with yield traits in the Indian tropical tasar silkworm Antheraea mylitta drury

The tropical tasar silkworm, Antheraea mylitta, is a semi-domesticated vanya silk-producing insect of high economic importance. To date, no molecular marker associated with cocoon and shell weights has been identified in this species. In this report, we identified a randomly amplified polymorphic DNA (RAPD) marker and examined its inheritance, and also developed a stable diagnostic sequence-characterized amplified region (SCAR) marker. Silkworms were divided into groups with high (HCSW) and low (LCSW) cocoon and shell weights, and the F₂ progeny of a cross between these two groups were obtained. DNA from these silkworms was screened by PCR using 34 random primers and the resulting RAPD fragments were used for cluster analysis and discriminant function analysis (DFA). The clustering pattern in a UPGMA-based dendogram and DFA clearly distinguished the HCSW and LCSW groups. Multiple regression analysis identified five markers associated with cocoon and shell weights. The marker OPW16_{905 bp} showed the most significant association with cocoon and shell weights, and its inheritance was confirmed in F₂ progeny. Cloning and sequencing of this 905 bp fragment showed 88% identity between its 134 nucleotides and the Bmc-1/Yamato-like retroposon of A. mylitta. This marker was further converted into a diagnostic SCAR marker (SCOPW 16_{826 bp}). The SCAR marker developed here may be useful in identifying the right parental stock of tasar silk-worms for high cocoon and shell weights in breeding programs designed to enhance the productivity of tasar silk.     

The dietary fatty acid 10E12Z-CLA induces epiregulin expression through COX-2 dependent PGF(2α) synthesis in adipocytes

Conjugated linoleic acids (CLAs) are a group of dietary fatty acids that are widely marketed as weight loss supplements. The isomer responsible for this effect is the trans-10, cis-12 CLA (10E12Z-CLA) isomer. 10E12Z-CLA treatment during differentiation of 3T3-L1 adipocytes induces expression of prostaglandin-endoperoxide synthase-2 (Cyclooxygenase-2; COX-2). This work demonstrates that COX-2 is also induced in fully differentiated 3T3-L1 adipocytes after a single treatment of 10E12Z-CLA at both the mRNA (20-40 fold) and protein level (7 fold). Furthermore, prostaglandin (PG)F(2α), but not PGE(2), is significantly increased 10 fold. In female BALB/c mice fed 0.5% 10E12Z-CLA for 10 days, COX-2 was induced in uterine adipose (2 fold). In vitro, pharmacological COX-2 inhibition did not block the effect of 10E12Z-CLA on adipocyte-specific gene expression although PGF(2α) was dose-dependently decreased. These studies demonstrate that PGF(2α) was not by itself responsible for the reduction in adipocyte character due to 10E12Z-CLA treatment. However, PGF(2α), either exogenously or endogenously in response to 10E12Z-CLA, increased the expression of the potent mitogen and epidermal growth factor (EGF) receptor (EGFR) ligand epiregulin in 3T3-L1 adipocytes. Blocking PGF(2α) signaling with the PGF(2α) receptor (FP) antagonist AL-8810 returned epiregulin mRNA levels back to baseline. Although this pathway is not directly responsible for adipocyte dependent gene expression, these results suggest that this signaling pathway may still have broad effect on the adipocyte and surrounding cells.     

Polymorphism in chemokine receptor genes and risk of acute myocardial infarction in North Indian population

Chemokines regulates the trafficking of leukocytes to the site of inflammation hence may be implicated in cardiac events. Currently no consistent effects have been revealed their role in acute myocardial infarction (MI). The aim of current study was to investigate the impact of human chemokine receptor genetic variants, CCR5-Δ32 insertion/deletion, CCR5-59029-A/G, CX3CR1-V249I and CX3CR1-T280 M on acute MI. 230 acute MI and 300 controls were examined. Patients carrying CCR5-Δ32 genotype were at three times higher risk of developing MI odds ratio (OR, 3.24, CI 1.127–9.356, P = 0.04). Significant association was found with risk of acute MI in recipients who possessed homozygous 59029-A allele (OR 1.47, CI 1.03–2.09, P = 0.03). While CX3CR1-I249 and M280 were found to be protective in MI patients with OR 0.46, CI 0.32–0.66, P < 0.0001 and OR 0.36, CI 0.24–0.55, P < 0.0001, respectively. It might be possible that risk of acute MI is associated with genetic variation in chemokine receptors, i.e., CCR5 and CX3CR1.     

Detection and imaging of the free radical DNA in cells--site-specific radical formation induced by Fenton chemistry and its repair in cellular DNA as seen by electron spin resonance, immuno-spin trapping and confocal microscopy

Oxidative stress-related damage to the DNA macromolecule produces lesions that are implicated in various diseases. To understand damage to DNA, it is important to study the free radical reactions causing the damage. Measurement of DNA damage has been a matter of debate as most of the available methods measure the end product of a sequence of events and provide limited information on the initial free radical formation. We report a measurement of free radical damage in DNA induced by a Cu(II)-H(2)O(2) oxidizing system using immuno-spin trapping supplemented with electron paramagnetic resonance. In this investigation, the short-lived radical generated is trapped by the spin trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO) immediately upon formation. The DMPO adduct formed is initially electron paramagnetic resonance active, but is subsequently oxidized to the stable nitrone adduct, which can be detected and visualized by immuno-spin trapping and has the potential to be further characterized by other analytical techniques. The radical was found to be located on the 2'-deoxyadenosine (dAdo) moiety of DNA. The nitrone adduct was repaired on a time scale consistent with DNA repair. In vivo experiments for the purpose of detecting DMPO-DNA nitrone adducts should be conducted over a range of time in order to avoid missing adducts due to the repair processes.     

Crystal structures of a type-1 ribosome inactivating protein from Momordica balsamina in the bound and unbound states

The ribosome inactivating proteins (RIPs) of type 1 are plant toxins that eliminate adenine base selectively from the single stranded loop of rRNA. We report six crystal structures, type 1 RIP from Momordica balsamina (A), three in complexed states with ribose (B), guanine (C) and adenine (D) and two structures of MbRIP-1 when crystallized with adenosine triphosphate (ATP) (E) and 2'-deoxyadenosine triphosphate (2'-dATP) (F). These were determined at 1.67?, 1.60?, 2.20?, 1.70?, 2.07? and 1.90? resolutions respectively. The structures contained, (A) unbound protein molecule, (B) one protein molecule and one ribose sugar, (C) one protein molecule and one guanine base, (D) one protein molecule and one adenine base, (E) one protein molecule and one ATP-product adenine molecule and (F) one protein molecule and one 2'-dATP-product adenine molecule. Three distinct conformations of the side chain of Tyr70 were observed with (i) χ(1)=-66°and χ(2)=165° in structures (A) and (B); (ii) χ(1)=-95° and χ(2)=70° in structures (C), (D) and (E); and (iii) χ(1)=-163° and χ(2)=87° in structure (F). The conformation of Tyr70 in (F) corresponds to the structure of a conformational intermediate. This is the first structure which demonstrates that the slow conversion of DNA substrates by RIPs can be trapped during crystallization.     

Lead induced antioxidant defense system in pigeon pea and its impact on yield and quality of seeds

Toxic effects of lead (>0.2 mM Pb) were measured in pigeon pea (Cajanus cajan Mill) cv. UPAS grown in sand culture as reduction in growth, yield, and quality of seeds. Leaves containing >38 μg g⁻¹ Pb showed oxidative damage as decrease in chlorophyll content and induction of antioxidants such as carotenoids, proline, and non-protein thiol contents with enhanced activities of SOD and peroxidase. At excess (>0.2 mM) Pb, accumulation of >1,000 μg Pb g⁻¹ root tissue was associated with increase in non-protein thiol content. It is concluded that inhibition in root-to-shoot translocation of Pb and induction in the level of proline, chloroplast pigments, and non-protein thiols and activities of antioxidant enzymes SOD and peroxidase at <0.2 mM Pb could have protected the pigeon pea plants from the deleterious effects of Pb. However, excess Pb at >0.2 mM showed a decline in yield, boldness, and quality of seeds despite the expression of an additional band each of Cu–Zn SOD and peroxidase isoform. The threshold of toxicity and toxicity values in pigeon pea leaves of plants exhibiting 10 and 33% yield depression at 27 days after metal supply were 27 and 56 μg Pb g⁻¹, respectively.     

Anti-neoplastic action of aspirin against a T-cell lymphoma involves an alteration in the tumour microenvironment and regulation of tumour cell survival

The present study explores the potential of the anti-neoplastic action of aspirin in a transplantable murine tumour model of a spontaneously originated T-cell lymphoma designated as Dalton's lymphoma. The antitumour action of aspirin administered to tumour-bearing mice through oral and/or intraperitoneal (intratumoral) routes was measured via estimation of survival of tumour-bearing mice, tumour cell viability, tumour progression and changes in the tumour microenvironment. Intratumour administration of aspirin examined to assess its therapeutic potential resulted in retardation of tumour progression in tumour-bearing mice. Oral administration of aspirin to mice as a prophylactic measure prior to tumour transplantation further primed the anti-neoplastic action of aspirin administered at the tumour site. The anti-neoplastic action of aspirin was associated with a decline in tumour cell survival, augmented induction of apoptosis and nuclear shrinkage. Tumour cells of aspirin-treated mice were found arrested in G0/G1 phase of the cell cycle and showed nuclear localization of cyclin B1. Intratumoral administration of aspirin was accompanied by alterations in the biophysical, biochemical and immunological composition of the tumour microenvironment with respect to pH, level of dissolved O2, glucose, lactate, nitric oxide, IFNγ (interferon γ), IL-4 (interleukin-4), IL-6 and IL-10, whereas the TGF-β (tumour growth factor-β) level was unaltered. Tumour cells obtained from aspirin-treated tumour-bearing mice demonstrated an altered expression of pH regulators monocarboxylate transporter-1 and V-ATPase along with alteration in the level of cell survival regulatory molecules such as survivin, vascular endothelial growth factor, heat-shock protein 70, glucose transporter-1, SOCS-5 (suppressor of cytokine signalling-5), HIF-1α (hypoxia-inducible factor-1α) and PUMA (p53 up-regulated modulator of apoptosis). The study demonstrates a possible indirect involvement of the tumour microenvironment in addition to a direct but limited anti-neoplastic action of aspirin in the retardation of tumour growth.     

The tomato genome: implications for plant breeding, genomics and evolution

ABSTRACT: The genome sequence of tomato (Solanum lycopersicum), one of the most important vegetable crops, has recently been decoded. We address implications of the tomato genome for plant breeding, genomics and evolutionary studies, and its potential to fuel future crop biology research.