Modification of Rifamycin Polyketide Backbone Leads to Improved Drug Activity against Rifampicin-resistant Mycobacterium tuberculosis

Rifamycin B, a product of Amycolatopsis mediterranei S699, is the precursor of clinically used antibiotics that are effective against tuberculosis, leprosy, and AIDS-related mycobacterial infections. However, prolonged usage of these antibiotics has resulted in the emergence of rifamycin-resistant strains of Mycobacterium tuberculosis. As part of our effort to generate better analogs of rifamycin, we substituted the acyltransferase domain of module 6 of rifamycin polyketide synthase with that of module 2 of rapamycin polyketide synthase. The resulting mutants (rifAT6::rapAT2) of A. mediterranei S699 produced new rifamycin analogs, 24-desmethylrifamycin B and 24-desmethylrifamycin SV, which contained modification in the polyketide backbone. 24-Desmethylrifamycin B was then converted to 24-desmethylrifamycin S, whose structure was confirmed by MS, NMR, and X-ray crystallography. Subsequently, 24-desmethylrifamycin S was converted to 24-desmethylrifampicin, which showed excellent antibacterial activity against several rifampicin-resistant M. tuberculosis strains.     

Rice Resistance to Planthoppers and Leafhoppers

For over 50 years, host-plant resistance has been regarded as an efficient method to reduce yield losses to rice caused by delphacid and cicadelid hoppers. Already a number of resistant rice varieties have been developed and deployed throughout Asia. To date, over 70 hopper resistance genes have been identified in rice; however, less than 10 genes have been deliberately introduced to commercial rice varieties. Currently, due to recent brown planthopper (Nilaparvata lugens [Stål]) and whitebacked planthopper (Sogatella furcifera [Horvath]) outbreaks occurring at an unprecedented scale, researchers are working toward a second generation of resistant varieties using newly identified gene loci and applying new molecular breeding methods. This paper reviews advances in the identification of resistance genes and QTLs against hoppers in rice. It collates all published information on resistance loci and QTLs against the major rice planthoppers and leafhoppers and presents information on gene locations, genetic markers, differential varieties, and wild rice species as sources of resistance. The review indicates that, whereas progress in the identification of genes has been rapid, considerable tidying of the information is required, especially regarding gene nomenclature and resistance spectra. Furthermore, sound information on gene functioning is almost completely lacking. However, hopper responses to resistance mechanisms are likely to be similar because a single phenotyping technique has been applied by most national and international breeding programs during germplasm screening. The review classifies genes occurring at two chromosome regions associated with several identified resistance loci and highlights these (Chr4S: BphR-R and Chr12L: BphR-R) as general stress response regions. The review calls for a greater diversity of phenotyping methods to enhance the durability of resistant varieties developed using marker-aided selection and emphasizes a need to anticipate the development of virulent hopper populations in response to the field deployment of genes.     

Population genetics and phylogenetics of DNA sequence variation at multiple loci within the Drosophila melanogaster species complex

Two regions of the genome, a 1-kbp portion of the zeste locus and a 1.1- kbp portion of the yolk protein 2 locus, were sequenced in six individuals from each of four species: Drosophila melanogaster, D. simulans, D. mauritiana, and D. sechellia. The species and strains were the same as those of a previous study of a 1.9-kbp region of the period locus. No evidence was found for recent balancing or directional selection or for the accumulation of selected differences between species. Yolk protein 2 has a high level of amino acid replacement variation and a low level of synonymous variation, while zeste has the opposite pattern. This contrast is consistent with information on gene function and patterns of codon bias. Polymorphism levels are consistent with a ranking of effective population sizes, from low to high, in the following order: D. sechellia, D. melanogaster, D.mauritiana, and D. simulans. The apparent species relationships are very similar to those suggested by the period locus study. In particular, D. simulans appears to be a large population that is still segregating variation that arose before the separation of D. mauritiana and D. sechellia. It is estimated that the separation of ancestral D. melanogaster from the other species occurred 2.5-3.4 Mya. The separations of D. sechellia and D. mauritiana from ancestral D. simulans appear to have occurred 0.58- 0.86 Mya, with D. mauritiana having diverged from ancestral D. simulans 0.1 Myr more recently than D. sechellia.      

A specific DNA sequence is required for high frequency of recombination in the ade6 gene of fission yeast.

The point mutation M26 in the ade6 gene of Schizosaccharomyces pombe increases recombination frequency by an order of magnitude in comparison with other mutations in the same gene. The hypothesis is tested that this hot spot of recombination requires a specific nucleotide sequence at the M26 site. The DNA sequence is altered systematically by in vitro mutagenesis, and the resulting sequences are introduced into the ade6 gene in vivo by gene replacement. It results that any change of the heptanucleotide ATGACGT leads to loss of high frequency of recombination. Thus this oligonucleotide sequence is necessary for high frequency of recombination, but it seems not to be sufficient.     

Effects of different mesenchymal stromal cell sources and delivery routes in experimental emphysema

We sought to assess whether the effects of mesenchymal stromal cells (MSC) on lung inflammation and remodeling in experimental emphysema would differ according to MSC source and administration route. Emphysema was induced in C57BL/6 mice by intratracheal (IT) administration of porcine pancreatic elastase (0.1 UI) weekly for 1 month. After the last elastase instillation, saline or MSCs (1×10⁵), isolated from either mouse bone marrow (BM), adipose tissue (AD) or lung tissue (L), were administered intravenously (IV) or IT. After 1 week, mice were euthanized. Regardless of administration route, MSCs from each source yielded: 1) decreased mean linear intercept, neutrophil infiltration, and cell apoptosis; 2) increased elastic fiber content; 3) reduced alveolar epithelial and endothelial cell damage; and 4) decreased keratinocyte-derived chemokine (KC, a mouse analog of interleukin-8) and transforming growth factor-β levels in lung tissue. In contrast with IV, IT MSC administration further reduced alveolar hyperinflation (BM-MSC) and collagen fiber content (BM-MSC and L-MSC). Intravenous administration of BM- and AD-MSCs reduced the number of M1 macrophages and pulmonary hypertension on echocardiography, while increasing vascular endothelial growth factor. Only BM-MSCs (IV > IT) increased the number of M2 macrophages. In conclusion, different MSC sources and administration routes variably reduced elastase-induced lung damage, but IV administration of BM-MSCs resulted in better cardiovascular function and change of the macrophage phenotype from M1 to M2.     

Phytotoxic effects of β-pinene on early growth and associated biochemical changes in rice

β-Pinene, an oxygenated monoterpene, is one of the major monoterpenes emitted into the atmosphere from forest areas and trees. Besides, it is a principal component of essential oils of a number of aromatic plants, which are involved in a variety of ecological interactions, including allelopathy, in the natural environment. However, studies pertaining to phytotoxicity and biochemical effect(s) of β-pinene are largely lacking. We investigated the effect of β-pinene (0.02, 0.04, 0.08, 0.20, 0.40 and 0.80 mg/ml) in a dose- and time-dependent manner on early seedling growth, dry weight accumulation, photosynthetic pigments and changes in macromolecule (protein and carbohydrate) content and activities of enzymes—proteases, α- and β-amylases, polyphenol oxidases and peroxidases- in rice (Oryza sativa) after 3rd, 5th and 7th day of exposure. β-pinene (≥0.04 mg/ml) significantly reduced the root (by 13–87%) and coleoptile (by 5–80%) length of rice. Exposure to β-pinene reduced total chlorophyll content in rice coleoptiles suggesting a negative impact on photosynthesis. The content of macromolecules (proteins and carbohydrates) enhanced significantly in response to β-pinene, whereas the activities of hydrolyzing enzymes—proteases, α-amylases, and β-amylases—declined (by 30–85, 26–84, 27–74%, respectively) in β-pinene-exposed seedlings. In contrast, the activities of peroxidases (POX) and polyphenol oxidases (PPO) enhanced significantly (by 16–152 and 53–290%, respectively) in rice roots in response to β-pinene in a dose- and time-dependent manner. Increased activities of POX and PPO indicate their involvement in providing protection and/or conferring resistance against β-pinene-induced stress. The study concludes that β-pinene inhibits the early growth of rice by altering the plant biochemical status and enhancing activities of POXs and PPOs involved in general plant defense.     

Dedifferentiation-mediated changes in transposition behavior make the Activator transposon an ideal tool for functional genomics in rice

There is an inverse relationship between the level of cytosine methylation in genomic DNA and the activity of plant transposable elements. Increased transpositional activity is seen during early plant development when genomic methylation patterns are first erased and then reset. Prolonging the period of hypomethylation might therefore result in an increased transposition frequency, which would be useful for rapid genome saturation in transposon-tagged plant lines. We tested this hypothesis using transgenic rice plants containing Activator (Ac) from maize. R₁ seeds from an Ac-tagged transgenic rice line were either directly germinated and grown to maturity, or induced to dedifferentiate in vitro, resulting in cell lines that were subsequently regenerated into multiple mature plants. Both populations were then analyzed for the presence, active reinsertion and amplification of Ac. Plants from each population showed excision-reinsertion events to both linked and unlinked sites. However, the frequency of transposition in plants regenerated from cell lines was more than nine-fold greater than that observed in plants germinated directly from seeds. Other aspects of transposon behavior were also markedly affected. For example, we observed a significantly larger proportion of transposition events to unlinked sites in cell line-derived plants. The tendency for Ac to insert into transcribed DNA was not affected by dedifferentiation. The differences in Ac activity coincided with a pronounced reduction in the level of genomic cytosine methylation in dedifferentiated cell cultures. We used the differential transposon behavior induced by dedifferentiation in the cell-line derived population for direct applications in functional genomics and validated the approach by recovering Ac insertions in a number of genes. Our results demonstrate that obtaining multiple Ac insertions is useful for functional annotation of the rice genome.These authors contributed equally to the work     

Status, invasiveness and environmental threats of three tropical American invasive weeds (Parthenium hysterophorus L., Ageratum conyzoides L., Lantana camara L.) in India

Invasive weeds have threatened the integrity of ecosystems throughout the world. They affect not only the species diversity of native areas but also their biological integrity. In India, a number of invasive exotic weeds have been reported but some viz. Parthenium hysterophorus, Lantana camara and Ageratum conyzoides, especially those from tropical America are troublesome and have caused adverse ecological, economic and social impact. These weeds can be seen growing in different landscapes but are luxuriantly localized in unattended forests and cultivated areas. Parthenium hysterophorus (Asteraceae, commonly known as congress grass) is perhaps the most troublesome and noxious weed of urban and rural India. Besides rapidly colonizing areas replacing the native vegetation, it is also known to cause a number of human health problems such as skin allergy, rhinitis and irritation to eyes of the residents in the vicinity. Likewise, it causes fodder scarcity in addition to being unpalatable and toxic to livestock. Lantana camara (Verbenaceae), another serious tropical American pest, has encroached upon large areas of land, especially the forests where it has virtually replaced the forest floor vegetation and reduced tree growth. Also because of its bushy and spreading type of growth it obstructs forest operations. The third weed, Ageratum conyzoides (Asteraceae, Billy goat weed) has invaded agricultural fields. It interferes with crops and causes yield reductions of major staple crops of India. When it invades rangeland areas, it out competes native grasses causing scarcity of fodder. These weeds have similar growth strategies such as fast growth rates, short life-cycles, greater reproductive potential, high competitive abilities and allelopathy that make them successful invaders of native habitats. Mechanical, chemical, biological and cultural control tactics have failed individually, though integrated approaches combining all these approaches along with community participation and proper land management have been relatively successful. This paper presents various aspects of biology, ecology, hazards and control measures of these weeds.     

Atomic structure of the major capsid protein of rotavirus: implications for the architecture of the virion

The structural protein VP6 of rotavirus, an important pathogen responsible for severe gastroenteritis in children, forms the middle layer in the triple-layered viral capsid. Here we present the crystal structure of VP6 determined to 2 A resolution and describe its interactions with other capsid proteins by fitting the atomic model into electron cryomicroscopic reconstructions of viral particles. VP6, which forms a tight trimer, has two distinct domains: a distal beta-barrel domain and a proximal alpha-helical domain, which interact with the outer and inner layer of the virion, respectively. The overall fold is similar to that of protein VP7 from bluetongue virus, with the subunits wrapping about a central 3-fold axis. A distinguishing feature of the VP6 trimer is a central Zn(2+) ion located on the 3-fold molecular axis. The crude atomic model of the middle layer derived from the fit shows that quasi-equivalence is only partially obeyed by VP6 in the T = 13 middle layer and suggests a model for the assembly of the 260 VP6 trimers onto the T = 1 viral inner layer.