Integration of optogenetics with complementary methodologies in systems neuroscience

Modern optogenetics can be tuned to evoke activity that corresponds to naturally occurring local or global activity in timing, magnitude or individual-cell patterning. This outcome has been facilitated not only by the development of core features of optogenetics over the past 10 years (microbial-opsin variants, opsin-targeting strategies and light-targeting devices) but also by the recent integration of optogenetics with complementary technologies, spanning electrophysiology, activity imaging and anatomical methods for structural and molecular analysis. This integrated approach now supports optogenetic identification of the native, necessary and sufficient causal underpinnings of physiology and behaviour on acute or chronic timescales and across cellular, circuit-level or brain-wide spatial scales

The sucrose non-fermenting 1-related kinase 2 gene SAPK9 improves drought tolerance and grain yield in rice by modulating cellular osmotic potential, stomatal closure and stress-responsive gene expression

Abstract Background Family members of sucrose non-fermenting 1-related kinase 2 (SnRK2), being plant-specific serine/threonine protein kinases, constitute the central core of abscisic acid (ABA)-dependent and ABA-independent signaling pathways, and are key regulators of abiotic stress adaptation in plants. We report here the functional characterization of SAPK9 gene, one of the 10 SnRK2s of rice, through developing gain-of-function and loss-of-function phenotypes by transgenesis. Results The gene expression profiling revealed that the abundance of single gene-derived SAPK9 transcript was significantly higher in drought-tolerant rice genotypes than the drought-sensitive ones, and its expression was comparatively greater in reproductive stage than the vegetative stage. The highest expression of SAPK9 gene in drought-tolerant Oryza rufipogon prompted us to clone and characterise the CDS of this allele in details. The SAPK9 transcript expression was found to be highest in leaf and upregulated during drought stress and ABA treatment. In silico homology modelling of SAPK9 with Arabidopsis OST1 protein showed the bilobal kinase fold structure of SAPK9, which upon bacterial expression was able to phosphorylate itself, histone III and OsbZIP23 as substrates in vitro. Transgenic overexpression (OE) of SAPK9 CDS from O. rufipogon in a drought-sensitive indica rice genotype exhibited significantly improved drought tolerance in comparison to transgenic silencing (RNAi) lines and non-transgenic (NT) plants. In contrast to RNAi and NT plants, the enhanced drought tolerance of OE lines was concurrently supported by the upgraded physiological indices with respect to water retention capacity, soluble sugar and proline content, stomatal closure, membrane stability, and cellular detoxification. Upregulated transcript expressions of six ABA-dependent stress-responsive genes and increased sensitivity to exogenous ABA of OE lines indicate that the SAPK9 is a positive regulator of ABA-mediated stress signaling pathways in rice. The yield-related traits of OE lines were augmented significantly, which resulted from the highest percentage of fertile pollens in OE lines when compared with RNAi and NT plants. Conclusion The present study establishes the functional role of SAPK9 as transactivating kinase and potential transcriptional activator in drought stress adaptation of rice plant. The SAPK9 gene has potential usefulness in transgenic breeding for improving drought tolerance and grain yield in crop plants.http://deepblue.lib.umich.edu/bitstream/2027.42/134605/1/12870_2016_Article_845.pd

Timing Offset Calibration of CZTI instrument aboard ASTROSAT

The radio as well as the high energy emission mechanism in pulsars is yet not understood properly. A multi-wavelength study is likely to help in better understanding of such processes. The first Indian space-based observatory, ASTROSAT, has five instruments aboard, which cover the electromagnetic spectrum from infra-red (1300 $\AA$) to hard X-ray (380 KeV). Cadmium Zinc Telluride Imager (CZTI), one of the five instruments is a hard X-ray telescope functional over an energy range of 20-380 KeV. We aim to estimate the timing offset introduced in the data acquisition pipeline of the instrument, which will help in time alignment of high energy time series with those from two other ground-based observatories, viz. the Giant Meterwave Radio Telescope (GMRT) and the Ooty Radio Telescope (ORT). PSR B0531+21 is a well-studied pulsar with nearly aligned radio and hard X-ray pulse profiles. We use simultaneous observations of this pulsar with the ASTROSAT, the ORT and the GMRT. The pulsar was especially observed using the ORT with almost daily cadence to obtain good timing solutions. We also supplement the ORT data with archival FERMI data for estimation of timing noise. The timing offset of ASTROSAT instruments was estimated from fits to arrival time data at the ASTROSAT and the radio observatories. We estimate the offset between the GMRT and the ASTROSAT-CZTI to be -4716 $\pm$ 50 $\mu s$. The corresponding offset with the ORT was -29639 $\pm$ 50 $\mu s$. The offsets between the GMRT and Fermi-LAT -5368 $\pm$ 56 $\mu s$. (Abridged)Comment: 6 pages, 5 figures, 2 tables, Revised and Updated, accepted for publication in A&

Sigma factor n, liaison to an ntrC and rpoS dependent regulatory pathway controlling acid resistance and the LEE in enterohemorrhagic Escherichia coli

Enterohemorrhagic Escherichia coli (EHEC) is dependent on acid resistance for gastric passage and low oral infectious dose, and the locus of enterocyte effacement (LEE) for intestinal colonization. Mutation of rpoN, encoding sigma factor N (σ(N)), dramatically alters the growth-phase dependent regulation of both acid resistance and the LEE. This study reports on the determinants of σ(N)-directed acid resistance and LEE expression, and the underlying mechanism attributable to this phenotype. Glutamate-dependent acid resistance (GDAR) in TW14359ΔrpoN correlated with increased expression of the gadX-gadW regulatory circuit during exponential growth, whereas upregulation of arginine-dependent acid resistance (ADAR) genes adiA and adiC in TW14359ΔrpoN did not confer acid resistance by the ADAR mechanism. LEE regulatory (ler), structural (espA and cesT) and effector (tir) genes were downregulated in TW14359ΔrpoN, and mutation of rpoS encoding sigma factor 38 (σ(S)) in TW14359ΔrpoN restored acid resistance and LEE genes to WT levels. Stability, but not the absolute level, of σ(S) was increased in TW14359ΔrpoN; however, increased stability was not solely attributable to the GDAR and LEE expression phenotype. Complementation of TW14359ΔrpoN with a σ(N) allele that binds RNA polymerase (RNAP) but not DNA, did not restore WT levels of σ(S) stability, gadE, ler or GDAR, indicating a dependence on transcription from a σ(N) promoter(s) and not RNAP competition for the phenotype. Among a library of σ(N) enhancer binding protein mutants, only TW14359ΔntrC, inactivated for nitrogen regulatory protein NtrC, phenocopied TW14359ΔrpoN for σ(S) stability, GDAR and ler expression. The results of this study suggest that during exponential growth, NtrC-σ(N) regulate GDAR and LEE expression through downregulation of σ(S) at the post-translational level; likely by altering σ(S) stability or activity. The regulatory interplay between NtrC, other EBPs, and σ(N)-σ(S), represents a mechanism by which EHEC can coordinate GDAR, LEE expression and other cellular functions, with nitrogen availability and physiologic stimuli.Microbiology and Molecular Genetic

Pregnancy in two patients of Glanzmann’s thrombasthenia: a rare case report

Glanzmann’s thrombasthenia (GT) is inherited platelet disorder with an autosomal recessive mode of inheritance. Though, quantitatively normal, the aggregation ability of platelets is reduced in this condition. Pregnancy and delivery are rare in these patients and have been associated with a high risk of severe postpartum hemorrhage. We describe two GT here 1st case was a primigravida, who was diagnosed to have GT 11 yrs back and was admitted as a term pregnancy which was terminated by elective caesarean section and was was successfully managed by platelet transfusion. 2nd case was a 24 year old lady got diagnosed as GT during the evaluation of frequent mucocutaneous bleed. Her antenatal management was like that of normal pregnancy. She was hospitalized 6 weeks prior to expected date of delivery. Elective LSCS (Lower Segment Caesarean Section) was done and was managed with 4 units of single donor platelet (SDP) without any other complication

Development of Region Specific Hybrid Goat and their Performance Evaluation under High Altitude Condition

Goat meat (chevon) comprises an important source of protein to provide essential amino acids in addition to other meat and plant sources of proteins. Therefore, demands for chevon are huge from civil and defence sector in this region. However, there is limited availability of fresh tender chevon in Ladakh region round the year. Hence, there was a need of augmenting local availability of fresh goat meat by developing animal technology for fast growing region-specific crossbred goat for meat purpose that can efficiently perform under adverse climatic conditions prevailing in this region. The present crossbred goat was developed by using mixing genes of adaptive and meat traits through cross breeding between local goats (Changthangi and Gaddi breeds of goats) and Sirohi/Black Bengal goats. To develop this technology, we introduced Black Bengal and Sirohi from plain areas and native breed of goats viz. Gaddi and Changthangi goats for further adaptation and growth performance studies at Leh-Ladakh. After initial studies goats were divided into high altitude resistant/adapted and susceptible groups. High altitude resistant/adapted goats were taken for further cross breeding and pure breeding. All the kids produced out of this breeding were studied for physiological responses, growth performance, and blood biochemical parameters to know their adaptive and growth performance at high altitude. Crossbred kids of Sirohi ♂/Black Bengal ♂ X Changthangi ♀ had significantly (P<0.05) higher weight gain, adaptive physiological responses and blood biochemicals level as compared to exotic pure bred and other cross bred kids. These crossbred kids attained market weight faster than local as well as breeds from plain areas (Sirohi and Black Bengal goats). Average meat yield is 7-10 kg per adult crossbred goat if slaughtered at 9-12 month age. These cross bred (broiler goat) may be reared at Leh-Ladakh for meat purpose. Hence, this animal technology may help in increasing of fresh goat meat (chevon) supply to meet army’s and civil requirements in Leh-Ladakh

Exogenously scavenged and endogenously synthesized heme are differentially utilized by Mycobacterium tuberculosis

Heme is both an essential cofactor and an abundant source of nutritional iron for the human pathogen Mycobacterium tuberculosis. While heme is required for M. tuberculosis survival and virulence, it is also potentially cytotoxic. Since M. tuberculosis can both synthesize and take up heme, the de novo synthesis of heme and its acquisition from the host may need to be coordinated in order to mitigate heme toxicity. However, the mechanisms employed by M. tuberculosis to regulate heme uptake, synthesis, and bioavailability are poorly understood. By integrating ratiometric heme sensors with mycobacterial genetics, cell biology, and biochemistry, we determined that de novo-synthesized heme is more bioavailable than exogenously scavenged heme, and heme availability signals the downregulation of heme biosynthetic enzyme gene expression. Ablation of heme synthesis does not result in the upregulation of known heme import proteins. Moreover, we found that de novo heme synthesis is critical for survival from macrophage assault. Altogether, our data suggest that mycobacteria utilize heme from endogenous and exogenous sources differently and that targeting heme synthesis may be an effective therapeutic strategy to treat mycobacterial infections. IMPORTANCE Mycobacterium tuberculosis infects ~25% of the world's population and causes tuberculosis (TB), the second leading cause of death from infectious disease. Heme is an essential metabolite for M. tuberculosis, and targeting the unique heme biosynthetic pathway of M. tuberculosis could serve as an effective therapeutic strategy. However, since M. tuberculosis can both synthesize and scavenge heme, it was unclear if inhibiting heme synthesis alone could serve as a viable approach to suppress M. tuberculosis growth and virulence. The importance of this work lies in the development and application of genetically encoded fluorescent heme sensors to probe bioavailable heme in M. tuberculosis and the discovery that endogenously synthesized heme is more bioavailable than exogenously scavenged heme. Moreover, it was found that heme synthesis protected M. tuberculosis from macrophage killing, and bioavailable heme in M. tuberculosis is diminished during macrophage infection. Altogether, these findings suggest that targeting M. tuberculosis heme synthesis is an effective approach to combat M. tuberculosis infections.Microbiology and Molecular Genetic

In Situ X‑ray Scattering Reveals Coarsening Rates of Superlattices Self-Assembled from Electrostatically Stabilized Metal Nanocrystals Depend Nonmonotonically on Driving Force

Self-assembly of colloidal nanocrystals (NCs) into superlattices (SLs) is an appealing strategy to design hierarchically organized materials with promising functionalities. Mechanistic studies are still needed to uncover the design principles for SL self-assembly, but such studies have been difficult to perform due to the fast time and short length scales of NC systems. To address this challenge, we developed an apparatus to directly measure the evolving phases in situ and in real time of an electrostatically stabilized Au NC solution before, during, and after it is quenched to form SLs using small-angle X-ray scattering. By developing a quantitative model, we fit the time-dependent scattering patterns to obtain the phase diagram of the system and the kinetics of the colloidal and SL phases as a function of varying quench conditions. The extracted phase diagram is consistent with particles whose interactions are short in range relative to their diameter. We find the degree of SL order is primarily determined by fast (subsecond) initial nucleation and growth kinetics, while coarsening at later times depends nonmonotonically on the driving force for self-assembly. We validate these results by direct comparison with simulations and use them to suggest dynamic design principles to optimize the crystallinity within a finite time window. The combination of this measurement methodology, quantitative analysis, and simulation should be generalizable to elucidate and better control the microscopic self-assembly pathways of a wide range of bottom-up assembled systems and architectures