Use of leaf meters to provide an instantaneous assessment of leaf chlorophyll has become common, but calibration of meter output into direct units of leaf chlorophyll concentration has been difficult and an understanding of the relationship between these two parameters has remained elusive. We examined the correlation of soybean (Glycine max) and maize (Zea mays L.) leaf chlorophyll concentration, as measured by organic extraction and spectrophotometric analysis, with output (M) of the Minolta SPAD-502 leaf chlorophyll meter. The relationship is non-linear and can be described by the equation chlorophyll (μmol m(-2))=10((M0.265)), r (2)=0.94. Use of such an exponential equation is theoretically justified and forces a more appropriate fit to a limited data set than polynomial equations. The exact relationship will vary from meter to meter, but will be similar and can be readily determined by empirical methods. The ability to rapidly determine leaf chlorophyll concentrations by use of the calibration method reported herein should be useful in studies on photosynthesis and crop physiology.

The impact of neonicotinoid insecticides on insect pollinators is highly controversial. Sublethal concentrations alter the behaviour of social bees and reduce survival of entire colonies 1-3 . However, critics argue that the reported negative effects only arise from neonicotinoid concentrations that are greater than those found in the nectar and pollen of pesticide-treated plants 4 . Furthermore, it has been suggested that bees could choose to forage on other available flowers and hence avoid or dilute exposure 4,5 . Here, using a two-choice feeding assay, we show that the honeybee, Apis mellifera, and the buff-tailed bumblebee, Bombus terrestris, do not avoid nectar-relevant concentrations of three of the most commonly-used neonicotinoids, imidacloprid (IMD), thiamethoxam (TMX), and clothianidin (CLO) in food. Moreover, bees of both species prefer to eat more of sucrose solutions laced with IMD or TMX than sucrose alone. Stimulation with IMD, TMX, and CLO neither elicited spiking responses from gustatory neurons in the bees' mouthparts nor inhibited the responses of sucrose-sensitive neurons. Our data indicate that bees cannot taste neonicotinoids and are not repelled by them. Instead, bees preferred solutions containing IMD or TMX even though the consumption of these pesticides caused them to eat less food overall. This work shows that bees cannot control their exposure to neonicotinoids in food and implies that treating flowering crops with IMD and TMX presents a significant hazard to foraging bees.Determining the impacts of pesticides on pollinators is important to resolve for the future of world food security. Pollinating insects like bees increase the yields of human crops, but in doing so, are inadvertently exposed to pesticides in floral nectar and pollen 6,7 . Several studies have concluded that bees exposed to sublethal doses of neonicotinoid pesticides in food have difficulty learning floral traits, feeding, navigating, and foraging 2,3,[8][9][10][11] and have impaired motor function 12 . These changes in behaviour often lead to colony failure 2,3 . This body of work has galvanized public concern over bee welfare, and in 2013, led to a two-year ban on the use of the three most common neonicotinoids (IMD, TMX, CLO) on flowering crops by the European Union. The agricultural importance of these pesticides has motivated

The NOTCH1 receptor is cleaved within its extracellular domain by furin during its maturation, yielding two subunits that are held together noncovalently by a juxtamembrane heterodimerization (HD) domain. Normal NOTCH1 signaling is initiated by the binding of ligand to the extracellular subunit, which renders the transmembrane subunit susceptible to two successive cleavages within and C terminal to the heterodimerization domain, catalyzed by metalloproteases and ␥-secretase, respectively. Because mutations in the heterodimerization domain of NOTCH1 occur frequently in human T-cell acute lymphoblastic leukemia (T-ALL), we assessed the effect of 16 putative tumor-associated mutations on Notch1 signaling and HD domain stability. We show here that 15 of the 16 mutations activate canonical NOTCH1 signaling. Increases in signaling occur in a ligand-independent fashion, require ␥-secretase activity, and correlate with an increased susceptibility to cleavage by metalloproteases. The activating mutations cause soluble NOTCH1 heterodimers to dissociate more readily, either under native conditions (n ‫؍‬ 3) or in the presence of urea (n ‫؍‬ 11). One mutation, an insertion of 14 residues immediately N terminal to the metalloprotease cleavage site, increases metalloprotease sensitivity more than all others, despite a negligible effect on heterodimer stability by comparison, suggesting that the insertion may expose the S2 site by repositioning it relative to protective NOTCH1 ectodomain residues. Together, these studies show that leukemia-associated HD domain mutations render NOTCH1 sensitive to ligand-independent proteolytic activation through two distinct mechanisms.The development of multicellular organisms is orchestrated by a limited number of highly conserved signaling pathways. One such pathway involves NOTCH receptors and downstream mediators, which can variously regulate the specification of cell fate, proliferation, self-renewal, survival, and apoptosis in a dose-and context-dependent fashion (3,47).Like other members of the NOTCH receptor family, mammalian NOTCH1 is a large multimodular type I transmembrane glycoprotein (Fig. 1A). During maturation, NOTCH1 undergoes proteolytic processing by furin at a site termed S1 that lies ϳ70 amino acids external to the transmembrane domain (25), yielding two noncovalently associated extracellular (N EC ) and transmembrane (N TM ) subunits (6,25,37). N EC contains 36 N-terminal epidermal growth factor (EGF)-like repeats that participate in binding to ligands (23, 39, 51) and three iterated LIN-12/NOTCH repeats (LNR), which help to maintain NOTCH receptors in the "off" state prior to ligand binding (13,24,40). The association of N EC and N TM is mediated by sequences lying immediately N terminal (HD-N) and C terminal (HD-C) of site S1; together, these sequences constitute the NOTCH subunit association, or "heterodimerization" (HD) domain (40).Binding of ligands to N EC triggers two sequential proteolytic events within the N TM subunit at sites S2 and S3. S2 cleavage occurs just...

BackgroundNotch receptors are normally cleaved during maturation by a furin-like protease at an extracellular site termed S1, creating a heterodimer of non-covalently associated subunits. The S1 site lies within a key negative regulatory region (NRR) of the receptor, which contains three highly conserved Lin12/Notch repeats and a heterodimerization domain (HD) that interact to prevent premature signaling in the absence of ligands. Because the role of S1 cleavage in Notch signaling remains unresolved, we investigated the effect of S1 cleavage on the structure, surface trafficking and ligand-mediated activation of human Notch1 and Notch2, as well as on ligand-independent activation of Notch1 by mutations found in human leukemia.Principal FindingsThe X-ray structure of the Notch1 NRR after furin cleavage shows little change when compared with that of an engineered Notch1 NRR lacking the S1-cleavage loop. Likewise, NMR studies of the Notch2 HD domain show that the loop containing the S1 site can be removed or cleaved without causing a substantial change in its structure. However, Notch1 and Notch2 receptors engineered to resist S1 cleavage exhibit unexpected differences in surface delivery and signaling competence: S1-resistant Notch1 receptors exhibit decreased, but detectable, surface expression and ligand-mediated receptor activation, whereas S1-resistant Notch2 receptors are fully competent for cell surface delivery and for activation by ligands. Variable dependence on S1 cleavage also extends to T-ALL-associated NRR mutations, as common class 1 mutations display variable decrements in ligand-independent activation when introduced into furin-resistant receptors, whereas a class 2 mutation exhibits increased signaling activity.Conclusions/SignificanceS1 cleavage has distinct effects on the surface expression of Notch1 and Notch2, but is not generally required for physiologic or pathophysiologic activation of Notch proteins. These findings are consistent with models for receptor activation in which ligand-binding or T-ALL-associated mutations lead to conformational changes of the NRR that permit metalloprotease cleavage.

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This study investigated relations among preschoolers' secure-base behavior with mother at home, stressful family conditions, and qualities of developing relationships with peers and teachers in preschool. Preschool-age children (N = 94; 51 boys and 43 girls) and their mothers were observed during multiple home visits, and their security of attachment was assessed with the Attachment Q-Set. Children in families with lower levels of stress had more secure attachment relationships with mothers. Both boys and girls who were less securely attached to their mothers expressed significantly more anger-aggression in preschool. For boys, higher levels of family stress were also significantly related to anger-aggression and to social competence in preschool. In addition, boys who were more securely attached to their mothers were more securely attached to their preschool teachers and were more popular with peers in preschool.

Summary Ins(3,4,5,6)P4 inhibits plasma membrane Cl− flux in secretory epithelia [1]. However, in most other mammalian cells, receptor-dependent elevation of Ins(3,4,5,6)P4 levels is an “orphan” response that lacks biological significance [2]. We set out to identify Cl− channel(s) and/or transporter(s) that are regulated by Ins(3,4,5,6)P4 in vivo. Several candidates [3-5] were excluded through biophysical criteria, electrophysiological analysis, and confocal immunofluorescence microscopy. Then, we heterologously expressed ClC-3 in the plasma membrane of HEK293-tsA201 cells; whole-cell patch-clamp analysis showed Ins(3,4,5,6)P4 to inhibit Cl− conductance through ClC-3. Next, we heterologously expressed ClC-3 in the early endosomal compartment of BHK cells; by fluorescence ratio imaging of endocytosed FITC-transferrin, we recorded intra-endosomal pH, an in situ biosensor for Cl− flux across endosomal membranes [6]. A cell-permeant, bioactivatable Ins(3,4,5,6)P4 analog elevated endosomal pH from 6.1 to 6.6, reflecting inhibition of ClC-3. Finally, Ins(3,4,5,6)P4 inhibited endogenous ClC-3 conductance in postsynaptic membranes of neonatal hippocampal neurones. Among other ClC-3 functions that could be regulated by Ins(3,4,5,6)P4 are tumor cell migration [7], apoptosis [8], and inflammatory responses [9]. Ins(3,4,5,6)P4 is a ubiquitous cellular signal with diverse biological actions.

The purpose of this article is to illuminate our troubles with, and troubling of, the trustworthiness dimension of balancing subjectivity and reflexivity, in qualitative research. This article evolved from debriefing sessions between three novice researchers working on a qualitative research study aimed at building understandings of the relational dynamics between adults with developmental disability diagnoses (ADevD) and their caregiving families. Following data collection, coauthors discussed interview experiences they had personally found challenging. These experiences constitute a point of departure for our examination of our researcher positions. We present a delineation of three research tensions, in the form of short "reflexive vignettes," each rooted in concern with possibly contradicting our goals of facilitating and expanding participant autonomy. We follow with recommendations about how, as researchers, our endeavor to understand participants with less conventional communication can be used to reflect and inform navigating difficulties universal to qualitative research.