Improvement of small seed for big nutritional feed

Exploding global population, rapid urbanization, salinization of soils, decreasing arable land availability, groundwater resources, and dynamic climatic conditions pose impending damage to our food security by reducing the grain quality and quantity. This issue is further compounded in arid and semi-arid regions due to the shortage of irrigation water and erratic rainfalls. Millets are gluten (a family of proteins)-free and cultivated all over the globe for human consumption, fuel, feed, and fodder. They provide nutritional security for the under- and malnourished. With the deployment of strategies like foliar spray, traditional/marker-assisted breeding, identification of candidate genes for the translocation of important minerals, and genome-editing technologies, it is now tenable to biofortify important millets. Since the bioavailability of iron and zinc has been proven in human trials, the challenge is to make such grains accessible. This review encompasses nutritional benefits, progress made, challenges being encountered, and prospects of enriching millet crops with essential minerals.     

Chronic Toxoplasma gondii in Nurr1-Null Heterozygous Mice Exacerbates Elevated Open Field Activity

Latent infection with Toxoplasma gondii is common in humans (approximately 30% of the global population) and is a significant risk factor for schizophrenia. Since prevalence of T. gondii infection is far greater than prevalence of schizophrenia (0.5-1%), genetic risk factors are likely also necessary to contribute to schizophrenia. To test this concept in an animal model, Nurr1-null heterozygous (+/-) mice and wild-type (+/+) mice were evaluate using an emergence test, activity in an open field and with a novel object, response to bobcat urine and prepulse inhibition of the acoustic startle response (PPI) prior to and 6 weeks after infection with T. gondii. In the emergence test, T. gondii infection significantly decreased the amount of time spent in the cylinder. Toxoplasma gondii infection significantly elevated open field activity in both +/+ and +/- mice but this increase was significantly exacerbated in +/- mice. T. gondii infection reduced PPI in male +/- mice but this was not statistically significant. Aversion to bobcat urine was abolished by T. gondii infection in +/+ mice. In female +/- mice, aversion to bobcat urine remained after T. gondii infection while the male +/- mice showed no aversion to bobcat urine. Antibody titers of infected mice were a critical variable associated with changes in open field activity, such that an inverted U shaped relationship existed between antibody titers and the percent change in open field activity with a significant increase in activity at low and medium antibody titers but no effect at high antibody titers. These data demonstrate that the Nurr1 +/- genotype predisposes mice to T. gondii-induced alterations in behaviors that involve dopamine neurotransmission and are associated with symptoms of schizophrenia. We propose that these alterations in murine behavior were due to further exacerbation of the altered dopamine neurotransmission in Nurr1 +/- mice.     

Laser Capture Microdissection - A Demonstration of the Isolation of Individual Dopamine Neurons and the Entire Ventral Tegmental Area

Laser capture microdissection (LCM) is used to isolate a concentrated population of individual cells or precise anatomical regions of tissue from tissue sections on a microscope slide. When combined with immunohistochemistry, LCM can be used to isolate individual cells types based on a specific protein marker. Here, the LCM technique is described for collecting a specific population of dopamine neurons directly labeled with tyrosine hydroxylase immunohistochemistry and for isolation of the dopamine neuron containing region of the ventral tegmental area using indirect tyrosine hydroxylase immunohistochemistry on a section adjacent to those used for LCM. An infrared (IR) capture laser is used to both dissect individual neurons as well as the ventral tegmental area off glass slides and onto an LCM cap for analysis. Complete dehydration of the tissue with 100% ethanol and xylene is critical. The combination of the IR capture laser and the ultraviolet (UV) cutting laser is used to isolate individual dopamine neurons or the ventral tegmental area when using PEN membrane slides. A PEN membrane slide has significant advantages over a glass slide as it offers better consistency in capturing and collecting cells, is faster collecting large pieces of tissue, is less reliant on dehydration and results in complete removal of the tissue from the slide. Although removal of large areas of tissue from a glass slide is feasible, it is considerably more time consuming and frequently leaves some residual tissue behind. Data shown here demonstrate that RNA of sufficient quantity and quality can be obtained using these procedures for quantitative PCR measurements. Although RNA and DNA are the most commonly isolated molecules from tissue and cells collected with LCM, isolation and measurement of microRNA, protein and epigenetic changes in DNA can also benefit from the enhanced anatomical and cellular resolution obtained using LCM.     

Isolation and functional characterization of three abiotic stress-inducible (Apx,Dhn and Hsc70) promoters from pearl millet (Pennisetum glaucum L.)

Molecular Biology Reports - Pearl millet is a C4 cereal crop that grows in arid and semi-arid climatic conditions with the remarkable abiotic stress tolerance. It contributed to the understanding...     

Antifungal benzo[b]thiophene 1,1-dioxide IMPDH inhibitors exhibit pan-assay interference (PAINS) profiles

Fungi cause serious life-threatening infections in immunocompromised individuals and current treatments are now complicated by toxicity issues and the emergence of drug resistant strains. Consequently, there is a need for development of new antifungal drugs. Inosine monophosphate dehydrogenase (IMPDH), a key component of the de novo purine biosynthetic pathway, is essential for growth and virulence of fungi and is a potential drug target. In this study, a high-throughput screen of 114,000 drug-like compounds against Cryptococcus neoformans IMPDH was performed. We identified three 3-((5-substituted)-1,3,4-oxadiazol-2-yl)thio benzo[b]thiophene 1,1-dioxides that inhibited Cryptococcus IMPDH and also possessed whole cell antifungal activity. Analogs were synthesized to explore the SAR of these hits. Modification of the fifth substituent on the 1,3,4-oxadiazole ring yielded compounds with nanomolar in vitro activity, but with associated cytotoxicity. In contrast, two analogs generated by substituting the 1,3,4-oxadiazole ring with imidazole and 1,2,4-triazole gave reduced IMPDH inhibition in vitro, but were not cytotoxic. During enzyme kinetic studies in the presence of DTT, nucleophilic attack of a free thiol occurred with the benzo[b]thiophene 1,1-dioxide. Two representative compounds with substitution at the 5 position of the 1,3,4-oxadiazole ring, showed mixed inhibition in the absence of DTT. Incubation of these compounds with Cryptococcus IMPDH followed by mass spectrometry analysis showed non-specific and covalent binding with IMPDH at multiple cysteine residues. These results support recent reports that the benzo[b]thiophene 1,1-dioxides moiety as PAINS (pan-assay interference compounds) contributor.     

Activity-Based Proteomic Profiling of Deubiquitinating Enzymes in Salmonella-Infected Macrophages Leads to Identification of Putative Function of UCH-L5 in Inflammasome Regulation

Although protein ubiquitination has been shown to regulate multiple processes during host response to Salmonella enterica serovar Typhimurium infection, specific functions of host deubiquitinating enzymes remain unknown in this bacterial infection. By using chemical proteomics approach, in which deubiquitinating enzymes were labeled by an active-site probe and analyzed by quantitative proteomics, we identified novel deubiquitinases in chicken macrophages based on their reactivity with the probe. Also, we detected down-regulation of UCH-L3, and USP4 as well as up-regulation of USP5 and UCH-L5 deubiquitinating enzymes in macrophages infected with Salmonella Typhimurium. We showed that decrease in either UCH-L5 activity, or in UCH-L5 protein amount in chicken and human macrophages infected or stimulated with LPS/nigericin, led to decreased IL-1β release. These data point towards a putative role of UCH-L5 in inflammasome regulation during Salmonella infection. Because inflammasome activation is important in innate resistance to these bacteria, one would expect that naturally occurring or therapeutically induced alteration in UCH-L5 activation would influence disease outcome and could represent a target for new therapeutic approaches.