In vitro organogenesis and plant regeneration of Thymus serpyllum L.: an important aromatic medicinal plant

An efficient in vitro propagation system has been developed for the rapid micropropagation of Thymus serpyllum L. (Banajwain), an aromatic medicinal herb from nodal explant on MS medium. Phenolic leaching and high rate of contamination was the most significant problem in establishing in vitro culture of Thymus serpyllum which was overcome by preparing explants in an antioxidant ascorbic acid (1000 ppm) at 6°C for 45 min and addition of the same antioxidant (50 mgl⁻¹) to the MS medium. The frequency of shoot production was influenced by different cytokinins (Kn, BAP, and Kn + BAP) and 95.56% shoot induction was observed when MS medium was supplemented with 1.0 + 2.0 mgl⁻¹ (Kn + BAP). The maximum average number of shoots 16.93 ± 2.15 and average length (3.98 ± 0.55) was recorded when MS medium have 0.5 + 2.0 mgl⁻¹ (Kn + BAP). The in vitro regenerated microshoots were rooted on MS and half strength MS medium and there was significant difference in root induction on both media under the influence of auxins (IAA, IBA, and NAA). The maximum average number (11.67 ± 3.03) and average root length (3.88 ± 0.71) was reported in half MS medium having 1.0 mgl⁻¹ IBA. The complete regenerated plantlets were acclimatized under growth chamber before transferring to the earthen pots and showed 90% survival.

Graphical abstract

     

Review of the biology,uses and conservation of the critically endangered endemic Himalayan species Nardostachys jatamansi (Caprifoliaceae)

Biodiversity and Conservation - The commercial demand for Nardostachys jatamansi in the global market has raised concern about its long-term sustainability. The genus Nardostachys is represented by...     

Rock phosphate solubilization by psychrotolerant Pseudomonas spp. and their effect on lentil growth and nutrient uptake under polyhouse conditions

Psychrotolerant Pseudomonas isolates (RT5RP2 and RT6RP) isolated from the rhizoplane of wild grass at 3,100 and 3,800 m above mean sea level, respectively, from Rudraprayag district of Uttarakhand (India), were found to solubilize Udaipur rock phosphate (URP). Both isolates grew at temperatures ranging from 4 to 30 °C. Kinetics of phosphate solubilization by the bacterial strains showed a nonlinear regression of the rate of P solubilization, which fitted best in the power model, and showed a declining trend across three different temperatures. Under pot culture conditions, bacterization of lentil seeds (cv. VL Masoor 507) with the psychrotolerant Pseudomonas strains when combined with URP as a sole source of phosphorus resulting in significant enhancement in P uptake of the plants, compared to the application of rock phosphate alone.     

Spatial distribution and regeneration of Quercus semecarpifolia and Quercus floribunda in a subalpine forest of western Himalaya, India

Studies were conducted on population status and regeneration of Quercus semecarpifolia and Quercus floribunda in a subalpine forest of western Himalaya. Saplings of Q. semecarpifolia belonging to the 51-100 cm size class and those of Q. floribunda belonging to the 101-150 cm size class were well represented in studied subalpine forests. Comparable seedling emergence and establishment of both species, either in the habitat dominated by same or different species seems to be an adaptational strategy of these species in subalpine forests. Seedling to tree ratio for Q. semecarpifolia was higher in the stand dominated by Q. floribunda, while comparable pattern was observed for Q. floribunda in the stand dominated by Q. semecarpifolia. Establishment of higher percent of seedlings of one species in the habitats dominated by another species of Quercus appears to indicate an oligarchic nature of oaks in subalpine forests. This study describes regeneration strategies, and spatial distribution of these species in a subalpine forest of Uttarakhand.     

Global Wild Annual Lens Collection: A Potential Resource for Lentil Genetic Base Broadening and Yield Enhancement

Crop wild relatives (CWRs) are invaluable gene sources for various traits of interest, yet these potential resources are themselves increasingly threatened by the impact of climate change as well as other anthropogenic and socio-economic factors. The prime goal of our research was to cover all aspects of wild Lens genetic resource management like species characterization, agro-morphological evaluation, diversity assessment, and development of representative sets for its enhanced utilization in lentil base broadening and yield improvement initiatives. We characterized and evaluated extensively, the global wild annual Lens taxa, originating from twenty seven counties under two agro-climatic conditions of India consecutively for three cropping seasons. Results on various qualitative and quantitative characters including two foliar diseases showed wide variations for almost all yield attributing traits including multiple disease resistance in the wild species, L. nigricans and L. ervoides accessions. The core set developed from the entire Lens taxa had maximum representation from Turkey and Syria, indicating rich diversity in accessions originating from these regions. Diversity analysis also indicated wide geographical variations across genepool as was reflected in the core set. Potential use of core set, as an initial starting material, for genetic base broadening of cultivated lentil was also suggested.     

Structural and mutational studies on substrate specificity and catalysis of Salmonella typhimurium D-cysteine desulfhydrase

Salmonella typhimurium DCyD (StDCyD) is a fold type II pyridoxal 5' phosphate (PLP)-dependent enzyme that catalyzes the degradation of D-Cys to H(2)S and pyruvate. It also efficiently degrades β-chloro-D-alanine (βCDA). D-Ser is a poor substrate while the enzyme is inactive with respect to L-Ser and 1-amino-1-carboxy cyclopropane (ACC). Here, we report the X-ray crystal structures of StDCyD and of crystals obtained in the presence of D-Cys, βCDA, ACC, D-Ser, L-Ser, D-cycloserine (DCS) and L-cycloserine (LCS) at resolutions ranging from 1.7 to 2.6 ?. The polypeptide fold of StDCyD consisting of a small domain (residues 48-161) and a large domain (residues 1-47 and 162-328) resembles other fold type II PLP dependent enzymes. The structures obtained in the presence of D-Cys and βCDA show the product, pyruvate, bound at a site 4.0-6.0 ? away from the active site. ACC forms an external aldimine complex while D- and L-Ser bind non-covalently suggesting that the reaction with these ligands is arrested at Cα proton abstraction and transimination steps, respectively. In the active site of StDCyD cocrystallized with DCS or LCS, electron density for a pyridoxamine phosphate (PMP) was observed. Crystals soaked in cocktail containing these ligands show density for PLP-cycloserine. Spectroscopic observations also suggest formation of PMP by the hydrolysis of cycloserines. Mutational studies suggest that Ser78 and Gln77 are key determinants of enzyme specificity and the phenolate of Tyr287 is responsible for Cα proton abstraction from D-Cys. Based on these studies, a probable mechanism for the degradation of D-Cys by StDCyD is proposed.     

Importance of tyrosine residues of Bacillus stearothermophilus serine hydroxymethyltransferase in cofactor binding and L-allo-Thr cleavage

Serine hydroxymethyltransferase (SHMT) from Bacillus stearothermophilus (bsSHMT) is a pyridoxal 5'-phosphate-dependent enzyme that catalyses the conversion of L-serine and tetrahydrofolate to glycine and 5,10-methylene tetrahydrofolate. In addition, the enzyme catalyses the tetrahydrofolate-independent cleavage of 3-hydroxy amino acids and transamination. In this article, we have examined the mechanism of the tetrahydrofolate-independent cleavage of 3-hydroxy amino acids by SHMT. The three-dimensional structure and biochemical properties of Y51F and Y61A bsSHMTs and their complexes with substrates, especially L-allo-Thr, show that the cleavage of 3-hydroxy amino acids could proceed via Calpha proton abstraction rather than hydroxyl proton removal. Both mutations result in a complete loss of tetrahydrofolate-dependent and tetrahydrofolate-independent activities. The mutation of Y51 to F strongly affects the binding of pyridoxal 5'-phosphate, possibly as a consequence of a change in the orientation of the phenyl ring in Y51F bsSHMT. The mutant enzyme could be completely reconstituted with pyridoxal 5'-phosphate. However, there was an alteration in the lambda max value of the internal aldimine (396 nm), a decrease in the rate of reduction with NaCNBH3 and a loss of the intermediate in the interaction with methoxyamine (MA). The mutation of Y61 to A results in the loss of interaction with Calpha and Cbeta of the substrates. X-Ray structure and visible CD studies show that the mutant is capable of forming an external aldimine. However, the formation of the quinonoid intermediate is hindered. It is suggested that Y61 is involved in the abstraction of the Calpha proton from 3-hydroxy amino acids. A new mechanism for the cleavage of 3-hydroxy amino acids via Calpha proton abstraction by SHMT is proposed.     

Vaccinia virus l1 protein is required for cell entry and membrane fusion

Genetic and biochemical studies have provided evidence for an entry/fusion complex (EFC) comprised of at least eight viral proteins (A16, A21, A28, G3, G9, H2, J5, and L5) that together with an associated protein (F9) participates in entry of vaccinia virus (VACV) into cells. The genes encoding these proteins are conserved in all poxviruses, are expressed late in infection, and are components of the mature virion membrane but are not required for viral morphogenesis. In addition, all but one component has intramolecular disulfides that are formed by the poxvirus cytoplasmic redox system. The L1 protein has each of the characteristics enumerated above except that it has been reported to be essential for virus assembly. To further investigate the role of L1, we constructed a recombinant VACV (vL1Ri) that inducibly expresses L1. In the absence of inducer, L1 synthesis was repressed and vL1Ri was unable to form plaques or produce infectious progeny. Unexpectedly, assembly and morphogenesis appeared normal and the noninfectious virus particles were indistinguishable from wild-type VACV as determined by transmission electron microscopy and analysis of the component polypeptides. Notably, the L1-deficient virions were able to attach to cells but the cores failed to penetrate into the cytoplasm. In addition, cells infected with vL1Ri in the absence of inducer did not form syncytia following brief low-pH treatment even though extracellular virus was produced. Coimmunoprecipitation experiments demonstrated that L1 interacted with the EFC and indirectly with F9, suggesting that L1 is an additional component of the viral entry apparatus.     

Epi p 1, an allergenic glycoprotein of Epicoccum purpurascens is a serine protease

Epicoccum purpurascens (EP) is a ubiquitous saprophytic mould, the inhalant spores and mycelia of which are responsible for respiratory allergic disorders in 5-7% of population worldwide. The diagnosis/therapy of these disorders caused by fungi involves the use of standardized and purified fungal extracts. A 33.5 kDa glycoprotein, Epi p 1 released histamine from whole blood cells of EP allergic patients at a concentration of 50-ng protein. The high specific IgE values detected in EP hypersensitive sera indicated that Epi p 1 is capable of mediating type I hypersensitive reaction in predisposed individuals. It also showed protease activity by virtue of its dose dependent cleavage of serine protease specific synthetic substrate, N-benzoyl arginine ethyl ester hydrochloride (BAEE). The serine protease nature of Epi p 1 was confirmed by its N-terminal sequence (ADG/FIVAVELD/STY) homology to a subtilisin like serine protease. The protease activity of Epi p 1 may be responsible for making its way into the system of pre-disposed individuals through epithelial cell detachment and the histamine releasing ability by cross-linking of IgE antibodies on cell surface is the cause of its allergenic nature.