An investigation into the role of Arabidopsis thaliana NAD metabolising enzymes in plant cellular stress responses

Adverse growing conditions resulting from abiotic stresses e.g. pathogen attack results in large losses in crop yields. Understanding and improving plants tolerance to an unfavourable environment remains one of the objectives in the study of plant biotechnology. One growing theory for reducing plant susceptibility to a broad range of stresses is modulation of cellular energy homeostasis. NAD+ (along with ATP) is the most important of cellular energy transducers in the form of a hydrogen ion donor and acceptor. However, a bigger picture is emerging as its role broadens to include a coenzyme, precursor for secondary messengers and a substrate for protein modifications. Three groups of proteins use NAD+ as a substrate, the NAD+ protein deacetylases, poly ADP polymerases and ADP ribose cyclases. Work on these groups of proteins in model organisms have identified their involvement in many biological roles including DNA repair, increasing longevity, initiating apoptosis and regulation of transcription. However, their role in plants is largely unknown. The aim of this study has been to identify the genes involved which use NAD+ and to further characterise their role in plant stress responses. A summary of the main results follows, a) AtPARG2 and sirtuin At5g55760 null lines were more sensitive to DNA damaging agents. b) AtPARG2 lines showed a disruption in circadian rhythm resulting in early flowering. c) Sirtuin At5g09230 null lines were more sensitive to UV-B exposure d) AtPARP3 was highly upregulated with exogenous Abscisic acid application

Site-Specific Incorporation of Unnatural Amino Acids into Receptors Expressed in Mammalian Cells

AbstractWe describe an approach to achieve unnatural amino acid incorporation into channels and receptors expressed in mammalian cells. We show that microelectroporation provides a general method to deliver DNA, mRNA, and tRNA simultaneously. In both CHO cells and cultured neurons, microelectroporation efficiently delivers an in vitro transcribed, serine amber suppressor tRNA, leading to nonsense suppression in a mutant EGFP gene. In CHO cells, both natural and unnatural amino acids chemically appended to a suppressor tRNA are site specifically incorporated into the nicotinic acetylcholine receptor (nAChR). Electrophysiology confirms the expected functional consequences of the unnatural residue. The microelectroporation strategy described here is more general, less tedious, and less damaging to mammalian neuronal and nonneuronal cells than previous approaches to nonsense suppression in small cells and provides the first example of unnatural amino acid incorporation in mammalian cells using chemically aminoacylated tRNA

A CONNECTION BETWEEN MORAL IMPERATIVE AND WOMEN’S LEADERSHIP?

This paper describes a qualitative study of two school districts’ efforts to support school leaders in developing the knowledge and skills to analyze, understand, and transform system inequities. The results suggest that, despite being armed with knowledge and skills, leaders may be reluctant to act even when doing so is within the scope of their authority. This work draws from Fullan’s (2003) concept of moral imperative and examines the connection between moral imperative and women’s ways of leading. The findings have implications for educational leadership programs that seek to prepare school leaders capable of challenging and changing inequitable systems

Obliquity Constraints on an Extrasolar Planetary-Mass Companion

We place the first constraints on the obliquity of a planetary-mass companion outside of the solar system. Our target is the directly imaged system 2MASS J01225093–2439505 (2M0122), which consists of a 120 Myr 0.4 M⊙ star hosting a 12–27 M_J companion at 50 au. We constrain all three of the system's angular-momentum vectors: how the companion spin axis, the stellar spin axis, and the orbit normal are inclined relative to our line of sight. To accomplish this, we measure projected rotation rates (v sin i) for both the star and the companion using new near-infrared high-resolution spectra with NIRSPEC at Keck Observatory. We combine these with a new stellar photometric rotation period from TESS and a published companion rotation period from Hubble Space Telescope to obtain spin-axis inclinations for both objects. We also fitted multiple epochs of astrometry, including a new observation with NIRC2/Keck, to measure 2M0122b's orbital inclination. The three line-of-sight inclinations place limits on the true de-projected companion obliquity and stellar obliquity. We find that while the stellar obliquity marginally prefers alignment, the companion obliquity tentatively favors misalignment. We evaluate possible origin scenarios. While collisions, secular spin–orbit resonances, and Kozai–Lidov oscillations are unlikely, formation by gravitational instability in a gravito-turbulent disk—the scenario favored for brown dwarf companions to stars—appears promising

Reversible DNA micro-patterning using the fluorous effect

We describe a new method for the immobilisation of DNA into defined patterns with sub-micron resolution, using the fluorous effect. The method is fully reversible via a simple solvent wash, allowing the patterning, regeneration and re-patterning of surfaces with no degradation in binding efficiency following multiple removal/attachment cycles of different DNA sequences

Enhanced differentiation of retinal progenitor cells using microfabricated topographical cues

Due to the retina’s inability to replace photoreceptors lost during retinal degeneration, significant interest has been placed in methods to implant replacement cells. Polymer scaffolds are increasingly being studied as vehicles for cellular delivery to degenerated retinas. Previously, we fabricated poly(methyl methacrylate) thin film scaffolds that increased survival and integration of implanted retinal progenitor cells (RPCs). Additionally, these scaffolds minimized the trauma and cellular response associated with implantation of foreign bodies into mouse eyes. Here, we demonstrate that biodegradable polycaprolactone (PCL) thin film scaffolds can be fabricated with integrated microtopography. Microfabricated topography in a PCL thin film enhanced the attachment and organization of RPCs compared to unstructured surfaces. Using real-time quantitative polymerase chain reaction we also observed that attachment to microtopography induced cellular differentiation. RPCs grown on PCL thin films exhibited an increase in gene expression for the photoreceptor markers recoverin and rhodopsin, an increase in the glial and Müller cell marker GFAP, and a decrease in SOX2 gene expression (a marker for undifferentiated progenitor cells) compared to cells grown on unmodified tissue culture polystyrene (TCPS)

Use of Consultants by U.S. Foundations: Results of a Foundation Center Survey

This article presents the results of a survey launched in January 2014 by Foundation Center, in collaboration with the National Network of Consultants to Grantmakers, examining use of consultants by community, corporate, and independent foundations whose annual giving totals at least $100,000. The survey asked funders to report whether they used consultants in the past two years and, if so, how frequently and for what purposes; they were also asked to report their level of satisfaction with consultants’ work. Funders that did not engage consultants in the last two years were asked why not. The survey also sought open-ended responses about working with consultants. The survey found widespread use of consultants among foundations. While the results of this study tend to emphasize the benefits – taking advantage of external expertise, allowing staff to stay focused on what they do best, bringing fresh or neutral perspectives to the work – respondents were also clear that working with consultants has its challenges

No difference in orbital parameters of RV-detected giant planets between 0.1 and 5 au in single vs multi-stellar systems

Our Keck/NIRC2 imaging survey searches for stellar companions around 144 systems with radial velocity (RV) detected giant planets to determine whether stellar binaries influence the planets' orbital parameters. This survey, the largest of its kind to date, finds eight confirmed binary systems and three confirmed triple systems. These include three new multi-stellar systems (HD 30856, HD 86081, and HD 207832) and three multi-stellar systems with newly confirmed common proper motion (HD 43691, HD 116029, and HD 164509). We combine these systems with seven RV planet-hosting multi-stellar systems from the literature in order to test for differences in the properties of planets with semimajor axes ranging between 0.1-5 au in single vs multi-stellar systems. We find no evidence that the presence or absence of stellar companions alters the distribution of planet properties in these systems. Although the observed stellar companions might influence the orbits of more distant planetary companions in these systems, our RV observations currently provide only weak constraints on the masses and orbital properties of planets beyond 5 au. In order to aid future efforts to characterize long period RV companions in these systems, we publish our contrast curves for all 144 targets. Using four years of astrometry for six hierarchical triple star systems hosting giant planets, we fit the orbits of the stellar companions in order to characterize the orbital architecture in these systems. We find that the orbital plane of the secondary and tertiary companions are inconsistent with an edge-on orbit in four out of six cases.Comment: 34 pages, 10 figures, 16 tables, including 4 tables in machine readable format and 7 tables with online supplemental dat

Characterisation of a pucBA deletion mutant from Rhodopseudomonas palustris lacking all but the pucBAd genes

Rhodopseudomonas palustris is a species of purple photosynthetic bacteria that has a multigene family of puc genes that encode the alpha and beta apoproteins, which form the LH2 complexes. A genetic dissection strategy has been adopted in order to try and understand which spectroscopic form of LH2 these different genes produce. This paper presents a characterisation of one of the deletion mutants generated in this program, the pucBAd only mutant. This mutant produces an unusual spectroscopic form of LH2 that only has a single large NIR absorption band at 800 nm. Spectroscopic and pigment analyses on this complex suggest that it has basically a similar overall structure as that of the wild-type HL LH2 complex. The mutant has the unique phenotype where the mutant LH2 complex is only produced when cells are grown at LL. At HL the mutant only produces the LH1-RC core complex