A communication network within the cytoplasmic domain of toll-like receptors has remained conserved during evolution

Toll/IL-1R (TIR) domain, that is, the cytoplasmic domain, in toll-like receptors (TLRs) from different species showed high sequence conservation in stretches spread across the surface as well as the core of the domain. To probe the structure–function significance of these residues, especially those coming from the core of TIR domains, we analyzed molecular dynamics trajectories of sequence similarity based models of human TIR domains. This study brought forth that N-terminal of the TIR domain simultaneously interacts with the flanking residues of the BB loop and central β-sheets. At the same time, residues of the central β-strands form favorable contacts with the DD loop and C-terminal, thus forming a two-way circuit between the N- and C-termini. In this work, the array of intradomain interactions is termed as communication network. Importantly, the “hubs” of this communication network were found to be conserved in all human TLRs. Earlier mutagenesis–function correlation work brought forth that certain mutations in the “core” of the TIR domain of TLR4 (e.g. in IFI767–769AAA and L815A) led to almost complete abrogation of signaling and reasoning for this dramatic loss-of-function has remained unclear, since these sites are not surface exposed. Using MD studies, we show here that this communication network gets disrupted in mutants of human TLR4 which were earlier reported to be functionally compromised. Extension of MD studies to heterodimer of TLR1/2 suggested that this evolutionarily conserved communication network senses the interactions formed upon dimerization and relays it to surfaces which are not involved in direct interdomain contacts.     

Mucuna pruriens seed extract reduces oxidative stress in nigrostriatal tissue and improves neurobehavioral activity in paraquat-induced Parkinsonian mouse model

Parkinson’s disease (PD) is a neurodegenerative disease which causes rigidity, resting tremor and postural instability. Treatment for this disease is still under investigation. Mucuna pruriens (L.), is a traditional herbal medicine, used in India since 1500 B.C., as a neuroprotective agent. In this present study, we evaluated the therapeutic effects of aqueous extract of M. pruriens (Mp) seed in Parkinsonian mouse model developed by chronic exposure to paraquat (PQ). Results of our study revealed that the nigrostriatal portion of Parkinsonian mouse brain showed significantly increased levels of nitrite, malondialdehyde (MDA) and reduced levels of catalase compared to the control. In the Parkinsonian mice hanging time was decreased, whereas narrow beam walk time and foot printing errors were increased.     

Monitoring protein aggregation during thermal unfolding in circular dichroism experiments

Thermal unfolding monitored by spectroscopy or calorimetry is widely used to determine protein stability. Equilibrium thermodynamic analysis of such unfolding is often hampered by its irreversibility, which usually results from aggregation of thermally denatured protein. In addition, heat-induced protein misfolding and aggregation often lead to formation of amyloid-like structures. We propose a convenient method to monitor in real time protein aggregation during thermal folding/ unfolding transition by recording turbidity or 90 degrees light scattering data in circular dichroism (CD) spectroscopic experiments. Since the measurements of turbidity and 90 degrees light scattering can be done simultaneously with far- or near-UV CD data collection, they require no additional time or sample and can be directly correlated with the protein conformational changes monitored by CD. The results can provide useful insights into the origins of irreversible conformational changes and test the linkage between protein unfolding or misfolding and aggregation in various macromolecular systems, including globular proteins and protein-lipid complexes described in this study, as well as a wide range of amyloid-forming proteins and peptides.     

A scalable approach for discovering conserved active subnetworks across species

Overlaying differential changes in gene expression on protein interaction networks has proven to be a useful approach to interpreting the cell's dynamic response to a changing environment. Despite successes in finding active subnetworks in the context of a single species, the idea of overlaying lists of differentially expressed genes on networks has not yet been extended to support the analysis of multiple species' interaction networks. To address this problem, we designed a scalable, cross-species network search algorithm, neXus (Network-cross(X)-species-Search), that discovers conserved, active subnetworks based on parallel differential expression studies in multiple species. Our approach leverages functional linkage networks, which provide more comprehensive coverage of functional relationships than physical interaction networks by combining heterogeneous types of genomic data. We applied our cross-species approach to identify conserved modules that are differentially active in stem cells relative to differentiated cells based on parallel gene expression studies and functional linkage networks from mouse and human. We find hundreds of conserved active subnetworks enriched for stem cell-associated functions such as cell cycle, DNA repair, and chromatin modification processes. Using a variation of this approach, we also find a number of species-specific networks, which likely reflect mechanisms of stem cell function that have diverged between mouse and human. We assess the statistical significance of the subnetworks by comparing them with subnetworks discovered on random permutations of the differential expression data. We also describe several case examples that illustrate the utility of comparative analysis of active subnetworks.     

LIBS-Based Detection of Antioxidant Elements in Seeds of <Emphasis Type="Italic">Emblica officinalis</Emphasis>

The aim of the study was to determine the effect of the elements of the extract of seed from Emblica officinalis on antioxidant enzymes and osmotic fragility of erythrocytes membrane in normal as well as streptozotocin-induced severely diabetic albino Wister rats. The results revealed that the untreated diabetic rats exhibited increase in oxidative stress as indicated by significantly diminished activities of free radical scavenging enzymes such as catalase (CAT) and superoxide dismutase (SOD) by 37.5% (p < 0.001) and 18.6% (p < 0.01), respectively. However, the E. officinalis seed extract treatment showed marked improvements in CAT and SOD activities by 47.09% (p < 0.001) and 21.61% (p < 0.001), respectively. The enhanced lipid peroxidation by 30.87% (p < 0.001) in erythrocytes of untreated diabetic rats was significantly accentuated in the extract treated animals by 23.72% (p < 0.001). The erythrocytes showed increased osmotic fragility due to diabetes in terms of hemolysis. It attained the normal level in diabetic treated group. The findings thus suggest that E. officinalis seed extract has the potential to be exploited as an agent to boost the antioxidant system in the diabetic animal model. Laser-induced breakdown spectroscopy has been used as an analytical tool to detect major and minor elements like Mg, Fe, Na, K, Zn, Ca, H, O, C, and N present in the extract. The higher concentration of Ca (II), Mg (II) and Fe (II) as reflected by their intensities are responsible for the antioxidant potential of E. officinalis.     

Bcl-2 Family Members and Functional Electron Transport Chain Regulate Oxygen Deprivation-Induced Cell Death

The mechanisms underlying cell death during oxygen deprivation are unknown. We report here a model for oxygen deprivation-induced apoptosis. The death observed during oxygen deprivation involves a decrease in the mitochondrial membrane potential, followed by the release of cytochrome c and the activation of caspase-9. Bcl-X(L) prevented oxygen deprivation-induced cell death by inhibiting the release of cytochrome c and caspase-9 activation. The ability of Bcl-X(L) to prevent cell death was dependent on allowing the import of glycolytic ATP into the mitochondria to generate an inner mitochondrial membrane potential through the F(1)F(0)-ATP synthase. In contrast, although activated Akt has been shown to inhibit apoptosis induced by a variety of apoptotic stimuli, it did not prevent cell death during oxygen deprivation. In addition to Bcl-X(L), cells devoid of mitochondrial DNA (rho degrees cells) that lack a functional electron transport chain were resistant to oxygen deprivation. Further, murine embryonic fibroblasts from bax(-/-) bak(-/-) mice did not die in response to oxygen deprivation. These data suggest that when subjected to oxygen deprivation, cells die as a result of an inability to maintain a mitochondrial membrane potential through the import of glycolytic ATP. Proapoptotic Bcl-2 family members and a functional electron transport chain are required to initiate cell death in response to oxygen deprivation.     

SHORT HYPOCOTYL IN WHITE LIGHT1, a serine-arginine-aspartate-rich protein in Arabidopsis, acts as a negative regulator of photomorphogenic growth

Light is an important factor for plant growth and development. We have identified and functionally characterized a regulatory gene SHORT HYPOCOTYL IN WHITE LIGHT1 (SHW1) involved in Arabidopsis (Arabidopsis thaliana) seedling development. SHW1 encodes a unique serine-arginine-aspartate-rich protein, which is constitutively localized in the nucleus of hypocotyl cells. Transgenic analyses have revealed that the expression of SHW1 is developmentally regulated and is closely associated with the photosynthetically active tissues. Genetic and molecular analyses suggest that SHW1 acts as a negative regulator of light-mediated inhibition of hypocotyl elongation, however, plays a positive regulatory role in light-regulated gene expression. The shw1 mutants also display shorter hypocotyl in dark, and analyses of shw1 cop1 double mutants reveal that SHW1 acts nonredundantly with COP1 to control hypocotyl elongation in the darkness. Taken together, this study provides evidences that SHW1 is a regulatory protein that is functionally interrelated to COP1 and plays dual but opposite regulatory roles in photomorphogenesis.