Cryptochromes: Photochemical and structural insight into magnetoreception

Cryptochromes (CRYs) function as blue light photoreceptors in diverse physiological processes in nearly all kingdoms of life. Over the past several decades, they have emerged as the most likely candidates for light‐dependent magnetoreception in animals, however, a long history of conflicts between in vitro photochemistry and in vivo behavioral data complicate validation of CRYs as a magnetosensor. In this review, we highlight the origins of conflicts regarding CRY photochemistry and signal transduction, and identify recent data that provides clarity on potential mechanisms of signal transduction in magnetoreception. The review primarily focuses on examining differences in photochemistry and signal transduction in plant and animal CRYs, and identifies potential modes of convergent evolution within these independent lineages that may identify conserved signaling pathways.     

Adhesion G protein-coupled receptor L3 gene variants: Statistically significant association observed in the male Indo-caucasoid Attention deficit hyperactivity disorder probands

Molecular Biology Reports - Primary symptoms of Attention Deficit Hyperactivity Disorder (ADHD) are age inappropriate inattention, hyperactivity and impulsivity. Caucasoid individuals showed...     

Mutational analysis of the SARS virus Nsp15 endoribonuclease: identification of residues affecting hexamer formation

The severe acute respiratory syndrome (SARS) coronavirus virus non-structural protein 15 is a Mn2+-dependent endoribonuclease with specificity for cleavage at uridylate residues. To better understand structural and functional characteristics of Nsp15, 22 mutant versions of Nsp15 were produced in Escherichia coli as His-tagged proteins and purified by metal-affinity and ion-exchange chromatography. Nineteen of the mutants were soluble and were analyzed for enzymatic activity. Six mutants, including four at the putative active site, were significantly reduced in endoribonuclease activity. Two of the inactive mutants had unusual secondary structures compared to the wild-type protein, as measured by circular dichroism spectroscopy. Gel-filtration analysis, velocity sedimentation ultracentrifugation, and native gradient pore electrophoresis all showed that the wild-type protein exists in an equilibrium between hexamers and monomers in solution, with hexamers dominating at micromolar protein concentration, while native gradient pore electrophoresis also revealed the presence of trimers. A mutant in the N terminus of Nsp15 was impaired in hexamer formation and had low endoribonuclease activity, suggesting that oligomerization is required for endoribonuclease activity. This idea was supported by titration experiments showing that enzyme activity was strongly concentration-dependent, indicating that oligomeric Nsp15 is the active form. Three-dimensional reconstruction of negatively stained single particles of Nsp15 viewed by transmission electron microscopic analysis suggested that the six subunits were arranged as a dimer of trimers with a number of cavities or channels that may constitute RNA binding sites.     

Evaluation of macrocyclic Grb2 SH2 domain-binding peptide mimetics prepared by ring-closing metathesis of C-terminal allylglycines with an N-terminal beta-vinyl-substituted phosphotyrosyl mimetic

Preferential binding of ligands to Grb2 SH2 domains in beta-bend conformations has made peptide cyclization a logical means of effecting affinity enhancement. This is based on the concept that constraint of open-chain sequences to bend geometries may reduce entropy penalties of binding. The current study extends this approach by undertaking ring-closing metathesis (RCM) macrocyclization between i and i+3 residues through a process involving allylglycines and beta-vinyl-functionalized residues. Ring closure in this fashion results in minimal macrocyclic tetrapeptide mimetics. The predominant effects of such macrocyclization on Grb2 SH2 domain binding affinity were increases in rates of association (from 7- to 16-fold) relative to an open-chain congener, while decreases in dissociation rates were less pronounced (approximately 2-fold). The significant increases in association rates were consistent with pre-ordering of solution conformations to near those required for binding. Data from NMR experiments and molecular modeling simulations were used to interpret the binding results. An understanding of the conformational consequences of such i to i+3 ring closure may facilitate its application to other systems where bend geometries are desired.     

Synthesis and anticancer activity of certain mononuclear Ru (II) complexes

Bis(1,10-phenanthroline/2,2'-bipyridine) ruthenium(II)complexes containing TCP, TTZ OPBI, and BTSC ligands (where, TCP = 1-thiocarbamoyl-3,5-diphenyl-2-pyrazoline, TTZ = 2-(3,5-diphenyl-4,5-dihydropyrazol-1-yl)-4-phenylthiazole, OPBI = 2-hydroxyphenyl benzimidazole and BTSC = benzoin thiosemicarbazone) have been prepared and characterized. The spectral data suggested that the ligands were coordinated with the metal through nitrogen, sulfur and oxygen atoms. The target complexes were tested in vivo for anticancer activity against transplantable murine tumor cell line, Ehrlich's Ascitic Carcinoma (EAC). All these complexes increased the life span of the EAC-bearing mice, decreased their tumor volume and viable ascitic cell count as well as improved Hb, RBC and WBC counts. These results suggest that the Ru(II) complexes exhibit significant antitumor activity in EAC-bearing mice. It was also observed that the ruthenium complexes protected red blood cells from 2,2'-azo-bis(2-methylpropionamidine) dihydrochloride (AAPH)- induced hemolysis. The inhibitory effect was dose-dependent at a concentration of 20-120 microg/ml.     

Cytomegalovirus capsid protease: biological substrates are cleaved more efficiently by full-length enzyme (pUL80a) than by the catalytic domain (assemblin)

We compared the full-length capsid maturational protease (pPR, pUL80a) of human cytomegalovirus with its proteolytic domain (assemblin) for the ability to cleave two biological substrates, and we found that pPR is more efficient with both. Affinity-purified, refolded enzymes and substrates were combined under defined reaction conditions, and cleavage was monitored and quantified following staining of the resulting electrophoretically separated fragments. The enzymes were stabilized against self-cleavage by a single point mutation in each cleavage site (ICRMT-pPR and IC-assemblin). The substrates were pPR itself, inactivated by replacing its catalytic nucleophile (S132A-pPR), and the sequence-related assembly protein precursor (pAP, pUL80.5). Our results showed that (i) ICRMT-pPR is 5- to 10-fold more efficient than assemblin for all cleavages measured (i.e., the M site of pAP and the M, R, and I sites of S132A-pPR). (ii) Cleavage of substrate S132A-pPR proceeded M>R>I for both enzymes. (iii) Na(2)SO(4) reduced M- and R-site cleavage efficiency by ICRMT-pPR, in contrast to its enhancing effect for both enzymes on I site and small peptide cleavage. (iv) Disrupting oligomerization of either the pPR enzyme or substrate by mutating Leu382 in the amino-conserved domain reduced cleavage efficiency two- to fourfold. (v) Finally, ICRMT-pPR mutants that include the amino-conserved domain, but terminate with Pro481 or Tyr469, retain the enzymatic characteristics that distinguish pPR from assemblin. These findings show that the scaffolding portion of pPR increases its enzymatic activity on biologically relevant protein substrates and provide an additional link between the structure of this essential viral enzyme and its biological mechanism.     

Root, shoot tissues of Brassica juncea and Cereal secale promote potato health

Brassica species are increasingly being used as cover crops to suppress soil-borne diseases in potato cropping systems. Experiments were conducted in controlled environments and in the field to evaluate the effects of cover crop root or shoot or a combination of root and shoot tissues on potato root and tuber health. In a lab assay we examined the extent to which volatile compounds released from tissues of two cover crop species, rye (Cereale secale L.) and oriental mustard (Brassica juncea L.), could inhibit mycelium growth of two important potato diseases, Rhizoctonia solani and Pythium ultimum. Twenty-four hours into the lab assay, volatile compounds from all residues suppressed fungal growth. After 48 h, marked suppression of hyphal growth continued in the presence of mustard residues but not in the presence of rye tissues or the control without tissues. A 75 L volume container experiment evaluated the effect of incorporating different quantities of mustard shoot and root tissues (none, comparable to field level and fourfold field level) into R. solani and P. ultimum infested soil on potato growth, root health and tuber disease. In the container study, incorporating mustard shoots at the highest dose increased potato yield by 54% and reduced disease rating to 2.3 compared to a severe rating of 4.4 in the control. In the field trial, potato growth, root health and tuber disease levels were evaluated in plots where disease management involved either incorporation of mustard or rye cover crop roots, shoots and whole plants (roots plus shoots) or standard farmer practice of a fumigated fallow as a control. White root tissue was used as a health indicator, and averaged 58 and 78% in the fumigated control and mustard cover crop treatments, respectively. The highest healthy root tissue status (91%) was recorded where whole plants of mustard were incorporated. In contrast to the visual assessment of root and tuber health, tuber yield in the field was not influenced by cover crop treatment. Across experiments, the incorporation of or exposure to whole mustard plants was consistently effective at suppressing soil-borne fungi and promoting healthy roots and tubers, especially at higher rates of biomass. Mustard should be managed so as to maximize incorporated biomass for effective biofumigation. Multipurpose management requiring removal of mustard shoots is incompatible with promoting potato rhizosphere health.     

Projecting future expansion of invasive species: comparing and improving methodologies for species distribution modeling

Modeling the distributions of species, especially of invasive species in non‐native ranges, involves multiple challenges. Here, we developed some novel approaches to species distribution modeling aimed at reducing the influences of such challenges and improving the realism of projections. We estimated species–environment relationships for Parthenium hysterophorus L. (Asteraceae) with four modeling methods run with multiple scenarios of (i) sources of occurrences and geographically isolated background ranges for absences, (ii) approaches to drawing background (absence) points, and (iii) alternate sets of predictor variables. We further tested various quantitative metrics of model evaluation against biological insight. Model projections were very sensitive to the choice of training dataset. Model accuracy was much improved using a global dataset for model training, rather than restricting data input to the species’ native range. AUC score was a poor metric for model evaluation and, if used alone, was not a useful criterion for assessing model performance. Projections away from the sampled space (i.e., into areas of potential future invasion) were very different depending on the modeling methods used, raising questions about the reliability of ensemble projections. Generalized linear models gave very unrealistic projections far away from the training region. Models that efficiently fit the dominant pattern, but exclude highly local patterns in the dataset and capture interactions as they appear in data (e.g., boosted regression trees), improved generalization of the models. Biological knowledge of the species and its distribution was important in refining choices about the best set of projections. A post hoc test conducted on a new Parthenium dataset from Nepal validated excellent predictive performance of our ‘best’ model. We showed that vast stretches of currently uninvaded geographic areas on multiple continents harbor highly suitable habitats for parthenium. However, discrepancies between model predictions and parthenium invasion in Australia indicate successful management for this globally significant weed.     

Analysis of p53 and NF-κB signaling in modulating the cardiomyocyte fate during hypertrophy

Cardiac hypertrophy leading to eventual heart failure is the most common cause of mortality throughout the world. The triggering mechanisms for cardiac hypertrophy are not clear but both apoptosis and cell proliferation have been reported in sections of failing hearts. In this study, we utilized both angiotensin II (AngII) treatment of cardiomyocytes and aortic ligation in rats (Rattus norvegicus, Wistar strain) for induction of hypertrophy to understand the cellular factors responsible for activation of apoptotic or anti-apoptotic pathway. Hypertrophy markers (ANF, β-MHC), apoptotic proteins (Bax, Bad, Fas, p53, caspase-3, PARP), and anti-apoptotic or cell proliferation marker proteins (Bcl2, NF-κB, Ki-67) were induced significantly during hypertrophy, both in vitro as well as in vivo. Co-localization of both active caspase-3 and Ki-67 was observed in hypertrophied myocytes. p53 and NF-κBp65 binding to co-activator p300 was also increased in AngII treated myocytes. Inhibition of p53 resulted in downregulation of apoptosis, NF-κB activation, and NF-κB-p300 binding; however, NF-κB inhibition did not inhibit apoptosis or p53-p300 binding. Blocking of either p53 or NF-κB by specific inhibitors resulted in decrease in cell proliferation and hypertrophy markers, suggesting that p53 initially binds to p300 and then this complex recruits NF-κB. Thus, these results indicate the crucial role of p53 in regulating both apoptotic and cell proliferation during hypertrophy.     

Charge Generation and Recombination in an Organic Solar Cell with Low Energetic Offsets

Organic bulk heterojunction (BHJ) solar cells require energetic offsets between the donor and acceptor to obtain high short‐circuit currents (J_SC) and fill factors (FF). However, it is necessary to reduce the energetic offsets to achieve high open‐circuit voltages (V_OC). Recently, reports have highlighted BHJ blends that are pushing at the accepted limits of energetic offsets necessary for high efficiency. Unfortunately, most of these BHJs have modest FF values. How the energetic offset impacts the solar cell characteristics thus remains poorly understood. Here, a comprehensive characterization of the losses in a polymer:fullerene BHJ blend, PIPCP:phenyl‐C61‐butyric acid methyl ester (PC₆₁BM), that achieves a high V_OC (0.9 V) with very low energy losses (E_loss = 0.52 eV) from the energy of absorbed photons, a respectable J_SC (13 mA cm^?2), but a limited FF (54%) is reported. Despite the low energetic offset, the system does not suffer from field‐dependent generation and instead it is characterized by very fast nongeminate recombination and the presence of shallow traps. The charge‐carrier losses are attributed to suboptimal morphology due to high miscibility between PIPCP and PC₆₁BM. These results hold promise that given the appropriate morphology, the J_SC, V_OC, and FF can all be improved, even with very low energetic offsets.