High-frequency regeneration via multiple shoot induction of an elite recalcitrant cotton (Gossypium hirsutum L. cv Narashima) by using embryo apex

Cotton (Gossypium hirsutum L.) is one of the most commercially important fiber crops in the world. Compared with other crops, cotton represents a recalcitrant species for regeneration protocols. The development of efficient and rapid regeneration protocol for elite Indian cotton variety could help improve the quality characteristics and biotic or abiotic stress tolerance. Here we report a novel regeneration protocol in Indian cotton cultivar Narashima. The maximum number of multiple shoots obtained was 16 per explants, performance which has never been achieved in any prior reports. The embryo apex explants were isolated from 2 d old in vitro growing seedlings. Explants were cultured on MS medium containing different plant growth regulator combinations in order to induce multiple shoots. Among the tested combinations, the 2 mg/l of 6-benzylaminopurine (BAP) and 2 mg/l kinetin (KIN) proved to be most suited for achieving the maximum number of multiple shoots. The elongation of multiple shoots was obtained in media supplemented with gibberellic acid (GA3). The regenerated plants were successfully hardened in earthen pots after adequate acclimatization. This method avoids callus tissue, the stage of regeneration which may lead to somaclonal variation. The important feature of the presented method is shortening of regeneration time, as well as the induction of a high number of multiple shoots per explants. The present protocol may provide an efficient and rapid regeneration tool for obtaining more stable transformants from embryo apex explants of Indian cotton cultivar Narashima.     

Ligand-Induced Protein Mobility in Complexes of Carbonic Anhydrase II and Benzenesulfonamides with Oligoglycine Chains

This paper describes a biophysical investigation of residual mobility in complexes of bovine carbonic anhydrase II (BCA) and para-substituted benzenesulfonamide ligands with chains of 1–5 glycine subunits, and explains the previously observed increase in entropy of binding with chain length. The reported results represent the first experimental demonstration that BCA is not the rigid, static globulin that has been typically assumed, but experiences structural fluctuations upon binding ligands. NMR studies with ¹⁵N-labeled ligands demonstrated that the first glycine subunit of the chain binds without stabilization or destabilization by the more distal subunits, and suggested that the other glycine subunits of the chain behave similarly. These data suggest that a model based on ligand mobility in the complex cannot explain the thermodynamic data. Hydrogen/deuterium exchange studies provided a global estimate of protein mobility and revealed that the number of exchanged hydrogens of BCA was higher when the protein was bound to a ligand with five glycine subunits than when bound to a ligand with only one subunit, and suggested a trend of increasing number of exchanged hydrogens with increasing chain length of the BCA-bound ligand, across the series. These data support the idea that the glycine chain destabilizes the structure of BCA in a length-dependent manner, causing an increase in BCA mobility. This study highlights the need to consider ligand-induced mobility of even “static” proteins in studies of protein-ligand binding, including rational ligand design approaches.     

TDP-43, an ALS Linked Protein,Regulates Fat Deposition and Glucose Homeostasis

The identification of proteins which determine fat and lean body mass composition is critical to better understanding and treating human obesity. TDP-43 is a well-conserved RNA-binding protein known to regulate alternative splicing and recently implicated in the pathogenesis of amyotrophic lateral sclerosis (ALS). While TDP-43 knockout mice show early embryonic lethality, post-natal conditional knockout mice show weight loss, fat depletion, and rapid death, suggesting an important role for TDP-43 in regulating energy metabolism. Here we report, that over-expression of TDP-43 in transgenic mice can result in a phenotype characterized by increased fat deposition and adipocyte hypertrophy. In addition, TDP-43 over-expression in skeletal muscle results in increased steady state levels of Tbc1d1, a RAB-GTPase activating protein involved in Glucose 4 transporter (Glut4) translocation. Skeletal muscle fibers isolated from TDP-43 transgenic mice show altered Glut4 translocation in response to insulin and impaired insulin mediated glucose uptake. These results indicate that levels of TDP-43 regulate body fat composition and glucose homeostasis in vivo.     

Enhanced N-metabolites,ABA and IAA-conjugate in anthers instigate heat sensitivity in spring wheat

Unraveling the metabolic and phytohormonal changes in anthers exposed to heat stress would help identify mechanisms regulating heat stress tolerance during the sensitive reproductive stage. Two spring wheat genotypes contrasting for heat tolerance were exposed to heat stress during heading in controlled environment chambers. Anthers were collected from main and primary spikes for metabolic and phytohormonal profiling. A significant reduction in seed set (38%), grain number (54%) and grain weight (52%) per plant was recorded in the sensitive (KSG1177) but not in the tolerant (KSG1214) genotype under heat stress compared to control. Anther metabolite accumulation did not vary quantitatively between main and primary spikes. Hierarchical clustering of the genotypes and treatments using metabolites and phytohormones revealed a distinct cluster for KSG1177 under heat stress from that of control and KSG1214. A significant increase in N-based amino acids, ABA, IAA-conjugate and a decrease in polyamines and organic acids were observed in wheat anthers exposed to heat stress. Unlike KSG1214, a significantly higher accumulation of amino acids, ABA and IAA-conjugate in anthers of the sensitive KSG1177 was recorded under heat stress. These findings provide the rationale and direction towards developing molecular markers for enhancing heat stress tolerance in wheat.     

Conformational insights into the inhibitory mechanism of phyto-compounds against Src kinase family members implicated in psoriasis

Abstract

Psoriasis is a chronic immune mediated disorder of the skin. There is growing evidence that the Src family tyrosine kinases (SFK) are highly upregulated in psoriasis. The SFK are the key components of the signaling pathways triggering cell growth and differentiation in addition to the immune cascades. In the current work, the interactions between SFK and selective phyto-compounds were studied using molecular docking approach. Based on the results of docking and binding energy calculations quercetin was identified as potential lead compound. To get a deeper insight into the binding of quercetin with the SFK, a combined molecular dynamics and binding free energy calculations were performed. The binding of quercetin disrupted the intra-molecular contacts making the SFK compact except Src kinase. The MM/PBSA free energy decomposition analysis highlighted the significance of hydrophobic and polar residues which are involved in the binding of quercetin. An experimental validation was carried out against the activated forms of Fyn, Lyn and Src kinases, the top three proteins which showed high preference for quercetin. The flow cytometry analysis showed that the expression levels of Fyn, Lyn and Src kinases were dramatically increased in HaCaT cells. However, the treatment of quercetin at the concentration of 51.65 µM for 24?h markedly decreased their expression in HaCaT cells. Besides, similar results were also observed when the HaCaT cells were treated with the kinase inhibitor Ponitinib (1.43 µM) for 24?h.

Communicated by Ramaswamy H. Sarma     

Factors,mechanisms and implications of chromatin condensation and chromosomal structural maintenance through the cell cycle

A series of well-orchestrated events help in the chromatin condensation and the formation of chromosomes. Apart from the formation of chromosomes, maintenance of their structure is important, especially for the cell division. The structural maintenance of chromosome (SMC) proteins, the non-SMC proteins and the SMC complexes are critical for the maintenance of chromosome structure. While condensins have roles for the DNA compaction, organization, and segregation, the cohesin functions in a cyclic manner through the cell cycle, as a “cohesin cycle.” Specific mechanisms maintain the architecture of the centromere, the kinetochore and the telomeres which are in tandem with the cell cycle checkpoints. The presence of chromosomal territories and compactness differences through the length of the chromosomes might have implications on selective susceptibility of specific chromosomes for induced genotoxicity.