Isolation and screening of potential actinobacteria for rapid composting of rice straw

Rice straw is produced as a by-product from rice cultivation, which is composed largely of lignocellulosic materials amenable to general biodegradation. Lignocellulolytic actinobacteria can be used as a potential agent for rapid composting of bulky rice straw. Twenty-five actinobacteria isolates were isolated from various in situ and in vitro rice straw compost sources. Isolates A2, A4, A7, A9 and A24 were selected through enzymatic degradation of starch, cellulose and lignin followed by the screening for their adaptability on rice straw powder amended media. The best adapted isolate (A7) was identified as Micromonospora carbonacea. It was able to degrade cellulose, hemicelluloses and carbon significantly (P ≤ 0.05) over the control. C/N ratio was reduced to 18.1 from an initial value of 29.3 in 6 weeks of composting thus having the potential to be used in large scale composting of rice straw.     

Caspase-2 Short Isoform Interacts with Membrane-Associated Cytoskeleton Proteins to Inhibit Apoptosis

Caspase-2 (casp-2) is the most conserved caspase across species, and is one of the initiator caspases activated by various stimuli. The casp-2 gene produces several alternative splicing isoforms. It is believed that the long isoform, casp-2L, promotes apoptosis, whereas the short isoform, casp-2S, inhibits apoptosis. The actual effect of casp-2S on apoptosis is still controversial, however, and the underlying mechanism for casp-2S-mediated apoptosis inhibition is unclear. Here, we analyzed the effects of casp-2S on DNA damage induced apoptosis through “gain-of-function” and “loss-of-function” strategies in ovarian cancer cell lines. We clearly demonstrated that the over-expression of casp-2S inhibited, and the knockdown of casp-2S promoted, the cisplatin-induced apoptosis of ovarian cancer cells. To explore the mechanism by which casp-2S mediates apoptosis inhibition, we analyzed the proteins which interact with casp-2S in cells by using immunoprecipitation (IP) and mass spectrometry. We have identified two cytoskeleton proteins, Fodrin and α-Actinin 4, which interact with FLAG-tagged casp-2S in HeLa cells and confirmed this interaction through reciprocal IP. We further demonstrated that casp-2S (i) is responsible for inhibiting DNA damage-induced cytoplasmic Fodrin cleavage independent of cellular p53 status, and (ii) prevents cisplatin-induced membrane blebbing. Taken together, our data suggests that casp-2S affects cellular apoptosis through its interaction with membrane-associated cytoskeletal Fodrin protein.     

Polygalacturonase activity and variation in ripening of papaya fruit with tissue depth and heat treatment

Effects of tissue position (viz. outer vs inner mesocarp) and heat treatment (48°C, 20 min) on variations in polygalacturonase (EC 3.2.1.15 and EC 3.2.1.67) activity and ripening of fruits of Carica papaya L. cv. Backcross Solo were investigated. Polygalacturonase activity increased during ripening concomitantly with an increase in tissue softness and soluble polyuronide level. Throughout ripening, inner mesocarp tissue was softer and contained higher polygalacturonase activity than outer mesocarp tissue. Titratable acidity as well as ß-galactosidase (EC 3.2.1.23) activity also increased during ripening; however, unlike polygalacturonase, their level or activity was lower in inner than in outer mesocarp. Ascorbic acid could partially account for the increase in titratable acidity during ripening but contributed very little to the differences in titratable acid levels between outer and inner mesocarp. Heat treatment had no effect on either fruit softness or titratable acidity, but it markedly reduced the increase in ascorbic acid and polygalacturonase activity during ripening. Ripening, as reflected by changes in tissue softness and polygalacturonase activity, progressed outwardly from the interior towards the exterior of the fruit. The effect of heat treatment in suppressing polygalacturonase activity was relatively greater in inner than in outer mesocarp, suggesting that sensitivity of the enzyme to heat treatment may vary with stage of ripeness of the tissue.     

Oil palm (Elaeis guineensis) protoplasts: isolation, culture and microcallus formation

Procedures are deseribed for the efficient isolation of protoplasts from a variety of oil palm (Elaeis guineensis Jacq.) tissues. Various factors including donor source, composition of enzyme mixture and culture medium affected the yield and viability of the protoplasts Polyembryogenic cultures of oil palm were the most suitable starting material in terms of yield, viability and metabolic competence. Pectolyase Y-23 in association with cellulase and hemicellulase was required for the efficient release of protoplasts from the oil palm tissues. Limited cell division to form microcallus was observed at very low frequency (<0.01%) when glutathione and catalase were incorporated in the culture medium.Abbreviations 2,4-d dichlorophenoxyacetic acid - DTT dithiothreitol - MES 2[N-morpholino] ethanesulphonic acid - NAA 1-naphthalene acetic acid - PVP polyvinylpyrrolidone     

A sequence specific endonuclease from Micrococcus radiodurans

A new sequence specific endonuclease, MraI has been purified from Micrococcus radiodurans. This enzyme cleaves bacteriophage λ DNA at three sites, adenovirus type 2 DNA at more than 12 sites and has a unique site on ΦX174 DNA. It has no sites on SV40, PM2 and pBR322 DNA. The three sites on phage λ DNA are different from those cleaved by SmaI, XmaI and XorII. The sites of cleavage are located at 0.424, 0.447 and 0.834 fractional lengths on the physical map of λ DNA. MraI is shown to be an isoschizomer of SacII and SstII recognizing the palindromic nucleotide sequence ′5-CCGC↓GG-3′. The enzyme shows an absolute requirement of Mg²⁺, but is active in the absence of added 2-mercaptoethanol. The enzyme shows activity at a broad range of temperature and pH with an optimum at 45°C and pH 7.0. MraI represents the first restriction enzyme from a bacterium whose DNA lacks modified methylated bases.     

Cryptic phenology in plants: Case studies,implications, and recommendations

Plant phenology—the timing of cyclic or recurrent biological events in plants—offers insight into the ecology, evolution, and seasonality of plant‐mediated ecosystem processes. Traditionally studied phenologies are readily apparent, such as flowering events, germination timing, and season‐initiating budbreak. However, a broad range of phenologies that are fundamental to the ecology and evolution of plants, and to global biogeochemical cycles and climate change predictions, have been neglected because they are “cryptic”—that is, hidden from view (e.g., root production) or difficult to distinguish and interpret based on common measurements at typical scales of examination (e.g., leaf turnover in evergreen forests). We illustrate how capturing cryptic phenology can advance scientific understanding with two case studies: wood phenology in a deciduous forest of the northeastern USA and leaf phenology in tropical evergreen forests of Amazonia. Drawing on these case studies and other literature, we argue that conceptualizing and characterizing cryptic plant phenology is needed for understanding and accurate prediction at many scales from organisms to ecosystems. We recommend avenues of empirical and modeling research to accelerate discovery of cryptic phenological patterns, to understand their causes and consequences, and to represent these processes in terrestrial biosphere models.     

Enzymatic characterization of Catalase from Bacillus anthracis and prediction of critical residues using information theoretic measure of Relative Entropy

In order to cope up with the reactive oxygen species (ROS) generated by host innate immune response, most of the intracellular organisms express Catalase for the enzymatic destruction/detoxification of hydrogen peroxide, to combat its deleterious effects. Catalase thus, scavenges ROS thereby playing a pivotal role in facilitating the survival of the pathogen within the host, and thus contributes to its pathogenesis. Bacillus anthracis harbors five isoforms of Catalase, but none of them has been studied so far. Thus, this study is the first attempt to delineate the biochemical and functional characteristics of one of the isoforms of Catalase (Cat1.4) of B. anthracis, followed by identification of residues critical for catalysis. The general strategy used, so far for mutational analysis in Catalases is structure based, i.e. the residues in the vicinity of heme were mutated to decipher the enzymatic mechanism. However, in the present study, protein sequence analysis was used for the prediction of catalytically important residues of Catalase. Essential measures were adopted to ensure the accuracy of predictions like after retrieval of well-annotated sequences from the database with EC 1.11.1.6, preprocessing was done to remove irrelevant sequences. The method used for multiple alignment of sequences, was guided by structural alignment and thereafter, an information theoretic measure, Relative Entropy was used for the critical residue prediction. By exploiting this strategy, we identified two previously known essential residues, H55 and Y338 in the active site which were demonstrated to be crucial for the activity. We also identified six novel crucial residues (Q332, Y117, H215, W257, N376 and H146) located distantly from the active site. Thus, the present study highlights the significance of this methodology to identify not only those crucial residues which lie in the active site of Catalase, but also the residues located distantly.     

Mutations in the chloroplast inner envelope protein TIC100 impair and repair chloroplast protein import and impact retrograde signaling

Chloroplast biogenesis requires synthesis of proteins in the nucleocytoplasm and the chloroplast itself. Nucleus-encoded chloroplast proteins are imported via multiprotein translocons in the organelle’s envelope membranes. Controversy exists around whether a 1-MDa complex comprising TIC20, TIC100, and other proteins constitutes the inner membrane TIC translocon. The Arabidopsis thaliana cue8 virescent mutant is broadly defective in plastid development. We identify CUE8 as TIC100. The tic100^cue8 mutant accumulates reduced levels of 1-MDa complex components and exhibits reduced import of two nucleus-encoded chloroplast proteins of different import profiles. A search for suppressors of tic100^cue8 identified a second mutation within the same gene, tic100^soh1, which rescues the visible, 1 MDa complex-subunit abundance, and chloroplast protein import phenotypes. tic100^soh1 retains but rapidly exits virescence and rescues the synthetic lethality of tic100^cue8 when retrograde signaling is impaired by a mutation in the GENOMES UNCOUPLED 1 gene. Alongside the strong virescence, changes in RNA editing and the presence of unimported precursor proteins show that a strong signaling response is triggered when TIC100 function is altered. Our results are consistent with a role for TIC100, and by extension the 1-MDa complex, in the chloroplast import of photosynthetic and nonphotosynthetic proteins, a process which initiates retrograde signaling.

Complementary mutations in TIC100 of the chloroplast inner envelope membrane cause reductions or corrective improvements in chloroplast protein import, and highlight a signaling role.

IN A NUTSHELLBackground: Plants harvest energy from the sun and CO₂ from the air and convert them into the energy-rich molecules they, and eventually us, are made of. Plants do this, photosynthesis, in bodies called chloroplasts inside their cells. Chloroplasts, made of protein and membrane material, were, before plants evolved, free-living bacteria, but the synthesis of most of their proteins occurs outside them, using information carried by the cell’s nuclear DNA, so most proteins have to be brought into developing chloroplasts, across the double membrane surrounding them, through dedicated, selective channels, formed by TOC (outer) and TIC (inner envelope) proteins. The identity of those channels matters as it helps determine versions of chloroplasts suited for particular environments. Which TIC proteins constitute the inner envelope channel has been a matter of controversy.Question: A mutant Arabidopsis plant called cue8 is slow-to-green (young leaves begin almost white) and shows delayed chloroplast and plant development. We looked for the molecular identity of the CUE8 gene. We also caused further mutations in this mutant and searched whether any corrected the defects in cue8.Findings: We found the mutated gene causing the cue8 defects is the TIC100 gene. This is one essential component of the “TIC 1-MDa complex,” one of the two versions of the TIC import complex under debate. That complex is made of several proteins, all present at reduced levels in cue8. In laboratory assays in which proteins are imported into isolated chloroplasts, cue8 performed worse than normal plants for a photosynthetic and a housekeeping chloroplast protein. A corrective, “suppressor” mutant was identified, and it carried a second mutation in TIC100, one physically complementary to the first one. Both the single and the double (suppressed) mutant still were slow-to-green, which evidences a signaling role for import defects to the nucleus, making photosynthetic genes active or not.Next steps: Surprisingly the grasses, including the cereals, have one core protein of the TIC 1 MDa complex but not the rest (including TIC100). We don’t know how their TIC channels operate. We also need to learn how the information on the defect in protein import, which occurs at the chloroplast envelope, is relayed to the cell’s nucleus (but we do have some clues).     

Chiral Imidazolidin-4-one with catalytic amount of Dicationic ionic liquid act as a recoverable and reusable Organocatalyst for asymmetric Diels-Alder reaction

Imidazolidin-4-one is used as a recoverable organocatalyst for the asymmetric Diels-Alder reaction in the presence of catalytic amount of dicationic ionic liquid and trifluoroacetic acid as a co-catalyst. The Diels-Alder reaction between model substrate cyclopentadiene and crotonaldehyde gave the product in 95% conversion and 87% ee of the endo-product. The catalyst was shown better reusability when the 20 mol% of dicationic ionic liquid was used and catalyst was reused upto 5 cycles, conversion remains upto 3 recycles but ee of endo- 9 was slightly droped.