Life Sciences

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Shiv Nadar University

Objectives: Stem cell factor receptor, c-kit, is considered to be the master signalling molecule of haematopoietic stem cells. It develops the orchestral pattern of haematopoietic cell lineages, seen by its varying degree of omnipresence in progenitors, lineage committed and mature cells. We have investigated the effect of over-expressing c-kit on early recovery of the haematopoietic compartment, in irradiated hosts.Materials and methods: Normal bone marrow cells (BMCs) were transfected with Kitwt (wild-type c-kit) or its variant Kitmu (asp814tyr) by electroporation. Lethally irradiated mice were transplanted with normal or transfected congeneic BMCs. The effect of ectopic expression of c-kit on haematopoietic cell recovery was determined by analysing donor-derived cells. Furthermore, effects of both types of c-kit over-expression on progenitor and lineage-committed cells were examined by flow cytometric analysis of Sca-1 and lineage-committed (Lin+) cells respectively.Results: Hyper-activating Kitmu significantly improved recovery of the haematopoietic system in irradiated hosts. In vivo results showed that the donor-derived c-kit+ cell population was increased to more than 3-fold in the case of Kitmu-transfected cells compared to normal and Kitwt over-expressing BMCs. In general, survival of progenitor and committed cell was improved in the Kitmu over-expressing system compared to the other two cohorts.Conclusion: These results suggest that recruitment of the hyper-activating variant of c-kit (Kitmu) lead to early recovery of the bone marrow of lethally irradiated mice. © 2010 Blackwell Publishing Ltd.

(SoNS) School of Natural Sciences

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Cell Proliferation

Foe turned friend: Multiple functional roles attributable to hyper-activating stem cell factor receptor mutant in regeneration of the haematopoietic cell compartment

The TIM23 complex mediates translocation of proteins across, and their lateral insertion into, the mitochondrial inner membrane. Translocation of proteins requires both the membrane-embedded core of the complex and its ATP-dependent import motor. Insertion of some proteins, however, occurs in the absence of ATP, questioning the need for the import motor during lateral insertion. We show here that the import motor associates with laterally inserted proteins even when its ATPase activity is not required. Furthermore, our results suggest a role for the import motor in lateral insertion. Thus, the import motor is involved in ATP-dependent translocation and ATP-independent lateral insertion. © 2011 European Molecular Biology Organization.

EMBO Reports

Role of the import motor in insertion of transmembrane segments by the mitochondrial TIM23 complex

Rev is an essential HIV-1 regulatory protein which binds to the Rev responsive element (RRE) present within the env gene of HIV-1 RNA genome. This binding facilitates the transport of the RNA to the cytoplasm, which in turn triggers the switch between viral latency and active viral replication. Essential components of this complex have been localized to a minimal arginine rich Rev peptide and stem IIB region of RRE. A synthetic peptide known as RSG-1.2 binds with high binding affinity and specificity to the RRE-IIB than the Rev peptide, however the thermodynamic basis of this specificity has not yet been addressed. The present study aims to probe the thermodynamic origin of this specificity of RSG-1.2 over Rev Peptide for RRE-IIB. The temperature dependent melting studies show that RSG-1.2 binding stabilizes the RRE structure significantly (ΔT m = 4.3°C), in contrast to Rev binding. Interestingly the thermodynamic signatures of the binding have also been found to be different for both the peptides. At pH 7.5, RSG-1.2 binds RRE-IIB with a K a = 16.2±0.6×10 7 M -1 where enthalpic change ΔH = -13.9±0.1 kcal/mol is the main driving force with limited unfavorable contribution from entropic change TΔS = -2.8±0.1 kcal/mol. A large part of ΔH may be due to specific stacking between U72 and Arg15. In contrast binding of Rev (K a = 3.1±0.4×10 7 M -1) is driven mainly by entropy (ΔH = 0 kcal/mol and TΔS = 10.2±0.2 kcal/mol) which arises from major conformational changes in the RNA upon binding. © 2011 Kumar et al.

PLoS ONE

Specificity of RSG-1.2 peptide binding to RRE-IIB RNA element of HIV-1 over rev peptide is mainly enthalpic in origin

In the past decade there has been growing interest in the investigation of plasma bubble (PB), associated irregularities, which often profoundly impact satellite communication and navigation systems and produce ionospheric scintillations. The basic mechanism of generation and development of equatorial spread F (ESF) is the gravitational Rayleigh-Taylor instability. The trans-ionospheric signal scintillation causes considerable communication hazards at a wide range of frequencies and hence is of much practical interest. In the present study we have investigated the effect of geomagnetic storm on the low latitude ionospheric scintillation for the 03 August 2010 storm at Varanasi (25.30°N, 82.99°E), India. © 2011 IEEE.

2011 30th URSI General Assembly and Scientific Symposium, URSIGASS 2011

Study of equatorial low-latitude ionospheric irregularities during a recent geomagnetic storm of 03 August 2010 using GPS receiver

The “trimpi event” consists of a sudden change in phase and amplitude of the VLF transmitter signal followed by a slow recovery to the initial levels as the ionization decays. We report here preliminary observations of phase and amplitude perturbations (trimpis) on VLF signals propagating in the Earth-ionosphere waveguide. Four VLF transmitter signals are monitored at our low latitude station Varanasi (L = 1.07) using SoftPAL Receiver. This paper deals with the current understanding of lightening discharge associated processes that leads to the changes in the characteristic of the waveguide and thus variation in the received amplitude and/or phase of the VLF transmission signals observed for the period August 25, 2009-December 25, 2009. Occurrence of Trimpi events at low L-values can be interpreted as precipitation of very high energetic electrons which are capable of producing perturbations during daylight, or as evidence of the ‘fast’ Trimpi mechanism which can dominate at low L-value. This wave induced particle precipitation is assumed to occur due to gyroresonance between whistler-mode waves and counter streaming electrons. © 2011 by World Scientific Publishing Co. Pte. Ltd. All rights reserved.

Advances in Geosciences: Volume 27: Solar Terrestrial (ST)

Trimpi events and other amplitude perturbations observed at Varanasi using SoftPAL receiver

Abstract Hypobaric hypoxia is encountered at high altitude. It has a deleterious effect on cognitive functions. An important cause of memory impairment at high altitude is the impairment of neurotransmission. The present study investigates the role of cholinergic markers in hypobaric hypoxia-induced memory impairment. Rats were exposed to hypobaric hypoxia at 6100 m for 7 days in a simulated-decompression chamber. Memory performance was assessed using the Morris water maze task. Cholinergic markers such as acetylcholine, acetylcholinesterase, choline acetyltransferase, α-7-nicotinic acetylcholine receptor and M 1 muscarinic acetylcholine receptor were also evaluated along with neuronal morphology and DNA fragmentation. We found impairment in memory function along with a decrease in acetylcholine levels, increase in acetylcholinesterase activity, down regulation of choline acetyltransferase, α-7-nicotinic acetylcholine receptor and M 1 muscarinic acetylcholine receptor. We also found that cellular damage is associated with a significant increase in DNA fragmentation. However, administration of acetylcholinesterase inhibitors, such as physostigmine and galantamine, resulted in amelioration of the hypobaric hypoxia induced deleterious effects. It improved acetylcholine level, decreased acetylcholinesterase activity and increased the synthesis of acetylcholine by increasing choline acetyltransferase activity. Also, the acetylcholinesterase inhibitors improved neuronal morphology, perhaps by increasing the expression of α-7-nicotinic acetylcholine receptor and by reducing the acetylcholinesterase level in the cortex and the hippocampus. Therefore, our results suggest cholinergic dysfunction is one of the mechanisms involved in hypobaric hypoxia-induced memory impairment and that acetylcholinesterase inhibitors were able to restore cholinergic function and thus improve memory function. © 2011 Elsevier B.V. All rights reserved.

European Journal of Pharmacology

Role of cholinergic markers on memory function of rats exposed to hypobaric hypoxia

A truncated (29 residues from the N-terminus) and N-terminal His-tagged form of SP-0149 from pneumococcal strain ATCC BAA-334 was overexpressed and purified to homogeneity using affinity and gel-filtration chromatography. Diffraction quality crystals were grown at 293 K using the hanging-drop vapour-diffusion technique. X-ray diffraction data were collected to 2.3 {\AA} resolution from a single-crystal that belonged to the orthorhombic space group P2 12 12 1 with the unit-cell parameters a = 54.56, b = 75.61, c = 75.52 {\AA}. The calculated values of the Matthews coefficient assuming one molecule (with calculated molecular weight of 30 400 Da) in the crystal asymmetric unit and the corresponding solvent content were 2.56 {\AA} 3 Da -1 and 52.0%, respectively. {\textcopyright} 2011 International Union of Crystallography All rights reserved.

Acta Crystallographica Section F: Structural Biology and Crystallization Communications

Cloning, overexpression, purification, crystallization and preliminary X-ray studies of SP-0149, the substrate binding protein of an ABC transporter from Streptococcus pneumoniae

A set of morphological traits and SSR markers were used to determine the genetic relationship among 12 elite thermosensitive genic male sterile (TGMS) lines developed at three different research institutions of India. Agro-morphological data recorded on 20 morphological traits revealed a wide base of genetic variation and a set of four morphological traits could distinguish most of the TGMS lines. Analysis with 30 SSR markers (20 EST-SSRs and 10 genomic SSRs) revealed 27 markers to be polymorphic, amplifying a total of 83 alleles. Each SSR marker amplified 2-6 alleles with an average of 2.76 alleles per marker and a PIC value varying from 0.54 to 0.96. Cluster analysis based on SSR and morphological data clearly differentiated the lines according to their source of origin. Correlation analysis between morphological and molecular data revealed a very poor association (r = 0.06), which could be attributed to selection pressure, genetic drift, sampling error and unknown relationship among related lines. The SSR markers discriminated the genotypes distinctly and quantified the genetic diversity precisely among the TGMS lines. Data on the yield per plant indicated that the genotypes grouping under a similar cluster showed same heterotic behaviour as compared to the genotypes from different clusters when crossed to similar pollinators. © 2011 Indian Academy of Sciences.

Journal of Genetics

Determination of genetic relationships among elite thermosensitive genic male sterile lines (TGMS) of rice (Oryza sativa L.) employing morphological and simple sequence repeat (SSR) markers

Marker assisted backcross breeding (MABB) provides a great opportunity for precise transfer of desirable donor segment by minimizing the linkage drag into a recurrent parent. In our lab, MABB was used for incorporating bacterial blight (BB) resistance genes (xa13 and Xa21) into the genetic background of Pusa Basmati1, which led to development of Improved Pusa Basmati 1 (Pusa 1460) as one of the first products of molecular breeding. Further, the parental lines of superfine grain aromatic rice hybrid Pusa RH 10 namely, Pusa 6A, Pusa 6B and PRR78 were improved for resistance to BB and blast by transferring genes xa13+Xa21 and Pi54 +Piz5, respectively. Presently, the pyramiding of genes for resistance to BB (xa13 and Xa21), blast (Piz5 and Pi54) and brown plant hopper (BPH; Bph 3, Bph 17, Bph 18, Bph20 and Bph 21) into Basmati rice varieties viz., Pusa Basmati 1121 and Pusa Basmati 6 is under way. In addition, a major QTL for salt tolerance (Saltol) is being transferred to Pusa Basmati 1121, which is widely grown in Haryana, the state having problem of salinity owing to underground brackish water. In order to develop genetically enhanced donor sources for resistance to biotic (BB, blast and BPH) and abiotic (salt tolerance, and phosphorus uptake) stresses in Basmati background, isogenic lines are being developed for major resistance genes/QTLs for respective stresses in the background of Pusa Basmati 1. Molecular mapping of fertility restorer gene(s) in Basmati restorer line PRR78 led to identification of two Rf gene linked molecular markers, RM258 and RM6100. Of these, RM6100 on validation in a set of rice germplasm showed 97.4% efficacy in identifying restorer lines from germplasm.QTL mapping using RIL population has unveiled several novel QTLs for grain and cooking quality traits. Molecular markers are also being routinely used for establishing variety/hybrids identity and authentication of genetic purity of hybrid seed lots.

Indian Journal of Genetics and Plant Breeding

Marker assisted selection: A paradigm shift in Basmati breeding

Background &amp; Aims: Hepatitis C virus (HCV) infection is characterized by lack of immune-mediated liver injury despite a high level of HCV replication during the incubation phase, which lasts about 8 weeks. We investigated whether this results from delayed recruitment of HCV-specific T cells and whether it facilitates HCV persistence. Methods: Six chimpanzees were infected with HCV; blood and liver samples were collected for 28 weeks and analyzed for immune cells and chemokines. Results: Two chimpanzees developed self-limited infections, whereas the remaining 4 developed chronic infections. Levels of the chemokines CXCL10, CXCL11, CCL4, and CCL5 increased in blood and liver samples from all chimpanzees within 1 month of HCV infection. Chemokine induction correlated with intrahepatic type I interferon (IFN) responses in vivo and was blocked by neutralizing antibodies against IFN-β in vitro. Despite the early-stage induction of chemokines, the intrahepatic lymphocytic infiltrate started to increase no earlier than 8 weeks after HCV infection, when HCV-specific, tetramer-positive CD8+ T cells appeared in the circulation. The HCV-specific CD8+ T cells expressed chemokine receptors when they were initially detected in blood samples, so they could be recruited to the liver as soon as they entered the circulation. Conclusions: Chemokines are induced during early stages of HCV infection, which requires a type I IFNmediated response. The delayed onset of acute hepatitis does not result from delayed recruitment of HCV-specific T cells, but could instead be related to a primary delay in the induction of HCV-specific T cells. Divergent outcomes occur without evident differences in chemokine induction and T-cell recruitment. © 2011 AGA Institute.