Miracle-dz/UniFAR-FTUs huggingface dataset:
Mutational Analyses of the Recombinant Globular Regions of Human C1q A, B, and C Chains Suggest an Essential Role for Arginine and Histidine Residues in the C1q-IgG Interaction1

The first step in the activation of the classical complement pathway by immune complexes involves the binding of the globular domain (gC1q) of C1q to the Fc regions of aggregated IgG or IgM. Each gC1q domain is a heterotrimer of the C-terminal halves of one A (ghA), one B (ghB), and one C (ghC) chain. Our recent studies have suggested a modular organization of gC1q, consistent with the view that ghA, ghB, and ghC are functionally autonomous modules and have distinct and differential ligand-binding properties. Although C1q binding sites on IgG have been previously identified, the complementary interacting sites on the gC1q domain have not been precisely defined. The availability of the recombinant constructs expressing ghA, ghB, and ghC has allowed us, for the first time, to engineer single-residue substitution mutations and identify residues on the gC1q domain, which are involved in the interaction between C1q and IgG. Because C1q is a charge pattern recognition molecule, we have sequentially targeted arginine and histidine residues in each chain. Consistent with previous chemical modification studies and the recent crystal structure of gC1q, our results support a central role for arginine and histidine residues, especially Arg114 and Arg129 of the ghB module, in the C1q-IgG interaction.
Sonographic findings in Beckwith–Wiedemann syndrome related to H19 hypermethylation

Beckwith–Wiedemann syndrome (BWS) is an overgrowth syndrome associated with congenital malformations and tumour predisposition. BWS results from variable mutations or epigenetic modifications of imprinted genes in the 11p15 chromosomal region. We present a fetus with mild general overgrowth and bilateral enlarged echogenic kidneys with loss of the corticomedullary differentiation in which prenatal diagnosis of BWS was suspected. The rest of the fetal anatomy and the amniotic fluid volume appeared normal. After termination of the pregnancy, molecular analysis confirmed the diagnosis of BWS by showing an isolated hypermethylation of the H19 gene. Copyright © 2004 John Wiley & Sons, Ltd.
Polysaccharide Compositions of Intervessel Pit Membranes Contribute to Pierce’s Disease Resistance of Grapevines1[OA]

Symptom development of Pierce’s disease (PD) in grapevine (Vitis vinifera) depends largely on the ability of the bacterium Xylella fastidiosa to use cell wall-degrading enzymes (CWDEs) to break up intervessel pit membranes (PMs) and spread through the vessel system. In this study, an immunohistochemical technique was developed to analyze pectic and hemicellulosic polysaccharides of intervessel PMs. Our results indicate that PMs of grapevine genotypes with different PD resistance differed in the composition and structure of homogalacturonans (HGs) and xyloglucans (XyGs), the potential targets of the pathogen’s CWDEs. The PMs of PD-resistant grapevine genotypes lacked fucosylated XyGs and weakly methyl-esterified HGs (ME-HGs), and contained a small amount of heavily ME-HGs. In contrast, PMs of PD-susceptible genotypes all had substantial amounts of fucosylated XyGs and weakly ME-HGs, but lacked heavily ME-HGs. The intervessel PM integrity and the pathogen’s distribution in Xylella-infected grapevines also showed differences among the genotypes. In pathogen-inoculated, PD-resistant genotypes PM integrity was well maintained and Xylella cells were only found close to the inoculation site. However, in inoculated PD-susceptible genotypes, PMs in the vessels associated with bacteria lost their integrity and the systemic presence of the X. fastidiosa pathogen was confirmed. Our analysis also provided a relatively clear understanding of the process by which intervessel PMs are degraded. All of these observations support the conclusion that weakly ME-HGs and fucosylated XyGs are substrates of the pathogen’s CWDEs and their presence in or absence from PMs may contribute to grapevine’s PD susceptibility.
Force per cross-sectional area from molecules to muscles: a general property of biological motors

We propose to formally extend the notion of specific tension, i.e. force per cross-sectional area—classically used for muscles, to quantify forces in molecular motors exerting various biological functions. In doing so, we review and compare the maximum tensions exerted by about 265 biological motors operated by about 150 species of different taxonomic groups. The motors considered range from single molecules and motile appendages of microorganisms to whole muscles of large animals. We show that specific tensions exerted by molecular and non-molecular motors follow similar statistical distributions, with in particular, similar medians and (logarithmic) means. Over the 1019 mass (M) range of the cell or body from which the motors are extracted, their specific tensions vary as Mα with α not significantly different from zero. The typical specific tension found in most motors is about 200 kPa, which generalizes to individual molecular motors and microorganisms a classical property of macroscopic muscles. We propose a basic order-of-magnitude interpretation of this result.
Structural rearrangement of CaMKIIα catalytic domains encodes activation

At its fundamental level, human memory is thought to occur at individual synaptic contact sites and manifest as persistent changes in synaptic efficacy. In digital electronics, the fundamental structure for implementing memory is the flip-flop switch, a circuit that can be triggered to flip between two stable states. Recently, crystals of Ca2+/calmodulin-dependent protein kinase IIα (CaMKIIα) catalytic domains, the enzymatic portion of a dodecameric holoenzyme involved in memory, were found to form dimers [Rosenberg OS, Deindl S, Sung RJ, Nairn AC, Kuriyan J (2005) Structure of the autoinhibited kinase domain of CaMKII and SAXS analysis of the holoenzyme. Cell 123:849–860]. Although the formation of dimers in the intact holoenzyme has not been established, several features of the crystal structure suggest that dimers could act as a synaptic switch. ATP-binding sites were occluded, and the T286 autophosphorylation site responsible for persistent kinase activation was buried. These features would act to stabilize an autoinhibited “paired”-enzyme state. Ca2+-calmodulin binding was postulated to trigger the formation of an active state with unpaired catalytic domains. This conformation would allow ATP access and expose T286, autophosphorylation of which would act to maintain the “unpaired” conformation. We used fluorescence anisotropy and FRET imaging of Venus-tagged CaMKIIα to test the hypothesis that neuronal CaMKIIα can flip between two stable conformations in living cells. Our data support the existence of catalytic domain pairs, and glutamate receptor activation in neurons triggered an increase in anisotropy consistent with a structural transition from a paired to unpaired conformation.
Kinetic Template‐Guided Tethering of Fragments

Birds of a tether: A tethering strategy for the site-directed discovery of low-molecular-weight fragments that bind weakly to defined protein surfaces is described. A solvent-exposed protein thiol captures acrylamide-modified fragments in a conjugate addition reaction that requires a template to produce a measureable quantity of protein-fragment adduct, which can be rapidly identified by mass spectrometry.
Evaluation of the BYG Carba Test, a New Electrochemical Assay for Rapid Laboratory Detection of Carbapenemase-Producing Enterobacteriaceae

ABSTRACT Accurate detection of carbapenemase-producing Enterobacteriaceae (CPE) constitutes a major laboratory diagnostic challenge. We evaluated an electrochemical technique (the BYG Carba test) which allows detection of CPE in less than 35 min. The BYG Carba test was first validated in triplicate against 57 collection isolates with previously characterized β-lactam resistance mechanisms (OXA-48, n = 12; KPC, n = 8; NDM, n = 8; VIM, n = 8; IMP, n = 3; GIM, n = 1; GES-6, n = 1; no carbapenemase, n = 16) and against a panel of 10 isolates obtained from the United Kingdom National External Quality Assessment Service (NEQAS). The test was then evaluated prospectively against 324 isolates referred to the national reference center for suspicion of CPE. The BYG Carba test results were compared with those obtained with the Carba NP test using multiplex PCR sequencing as the gold standard. Of the 57 collection and the 10 NEQAS isolates, all but one GES-6-producing isolate were correctly identified by the Carba BYG test. Among the 324 consecutive Enterobacteriaceae isolates tested prospectively, 146 were confirmed as noncarbapenemase producers by PCR while 178 harbored a carbapenemase gene (OXA-48, n = 117; KPC, n = 25; NDM, n = 23; and VIM, n = 13). Prospectively, in comparison with PCR results, the BYG Carba test displayed 95% sensitivity and 100% specificity versus 89% and 100%, respectively, for the Carba NP test. The BYG Carba test is a novel, rapid, and efficient assay based on an electro-active polymer biosensing technology discriminating between CPE and non-CPE. The precise electrochemical signal (electrochemical impedance variations) allows the establishment of real-time objective measurement and interpretation criteria which should facilitate the accreditation process of this technology.
Respiratory mechanics during exercise in endurance‐trained men and women

The purpose of this study was to compare the mechanics of breathing including the measurement of expiratory flow limitation, end‐expiratory lung volume, end‐inspiratory lung volume, and the work of breathing in endurance‐trained men (n= 8) and women (n= 10) during cycle exercise. Expiratory flow limitation was assessed by applying a negative expiratory pressure at the mouth. End‐expiratory lung volume and end‐inspiratory lung volume were determined by having subjects perform inspiratory capacity manoeuvres. Transpulmonary pressure, taken as the difference between oesophageal and airway opening pressure, was plotted against volume and integrated to determine the work of breathing. Expiratory flow limitation occurred in nine females (90%) and three males (43%) during the final stage of exercise. Females had a higher relative end‐expiratory lung volume (42 ± 8 versus 35 ± 5% forced vital capacity (FVC)) and end‐inspiratory lung volume (88 ± 5 versus 82 ± 7% FVC) compared to males at maximal exercise (P < 0.05). Women also had a higher work of breathing compared to men across a range of ventilations. On average, women had a work of breathing that was twice that of men at ventilations above 90 l min−1. These data suggest that expiratory flow limitation may be more common in females and that they experience greater relative increases in end‐expiratory lung volume and end‐inspiratory lung volume at maximal exercise compared to males. The higher work of breathing in women is probably attributed to their smaller lung volumes and smaller diameter airways. Collectively, these findings suggest that women utilize a greater majority of their ventilatory reserve compared to men and this is associated with a higher cost of breathing.
Changes in the concentrations of glucagon-like peptide-1(7–36)amide and gastric inhibitory polypeptide during the lactation cycle in goats

Plasma concentrations of glucagon-like peptide-1(7–36)amide (GLP) and gastric inhibitory polypeptide (GIP) were determined at fortnightly intervals for over a year throughout the pregnancy–lactation cycle of goats. Both GIP and GLP concentrations were elevated during lactation and fell rapidly when milk secretion was terminated. At the onset of lactation GLP concentrations rose rapidly whereas GIP concentrations showed a delayed response. GLP concentrations remained high throughout lactation but those of GIP declined linearly as milk yields fell. Serum insulin concentrations correlated positively with plasma glucose concentrations but not with either GIP or GLP concentrations. Negative correlations were found between serum insulin concentrations and milk yield and plasma non-esterified fatty acid concentrations. The results are consistent with plasma GIP and GLP concentrations being determined by other factors in addition to nutrient intake and absorption. Changes in GIP concentrations mirrored reported changes in the hypertrophy and atrophy of the intestine in ruminants while GLP concentrations may be more dependent on the neural and endocrine factors associated with lactation. The elevated concentrations of both peptides indicated a specific role in lactation independent of their normal anabolic and insulinotropic effects.
Allostatic and environmental load in toddlers predicts anxiety in preschool and kindergarten

Abstract Psychobiological models of allostatic load have delineated the effects of multiple processes that contribute to risk for psychopathology. This approach has been fruitful, but the interactive contributions of allostatic and environmental load remain understudied in early childhood. Because this developmental period encompasses the emergence of internalizing problems and biological sensitivity to early experiences, this is an important time to examine this process. In two studies, we examined allostatic and environmental load and links to subsequent internalizing and externalizing problems. Study 1 examined relations between load indices and maladjustment, concurrently and at multiple times between age 2 and kindergarten; Study 2 added more comprehensive risk indices in a sample following a group of highly fearful toddlers from 2 to 3 years of age. Results from both studies showed that increased allostatic load related to internalizing problems as environmental risk also increased. Study 2, in addition, showed that fearfulness interacted with allostatic and environmental load indices to predict greater anxiety among the fearful children who had high levels of allostatic and environmental load. Taken together, the findings support a model of risk for internalizing characterized by the interaction of biological and environmental stressors, and demonstrate the importance of considering individual differences and environmental context in applying models of allostatic load to developmental change in early childhood.
Phosphatidylinositol 3-Kinase-dependent Activation of Protein Kinase C-ζ in Bacterial Lipopolysaccharide-treated Human Monocytes*

The isoform identity of activated protein kinase C (PKC) and its regulation were investigated in bacterial lipopolysaccharide (LPS)-treated human monocytes. Resolution of detergent-soluble lysates prepared from LPS-treated, peripheral blood monocytes using Mono Q anion-exchange chromatography revealed two principal peaks of myelin basic protein kinase activity. Immunoblotting and immunoprecipitation with isoform-specific anti-PKC antibodies showed that the major and latest eluting peak is accounted for by PKC-ζ. In addition to primary monocytes, activation of PKC-ζ in response to LPS was also observed in the human promonocytic cell lines, U937 and THP-1. Consistent with its identity as PKC-ζ, the kinase did not depend upon the presence of lipids, Ca2+, or diacylglycerol for activity. In addition, the kinase phosphorylates peptide ε and myelin basic protein with equal efficiency but phosphorylates Kemptide and protamine sulfate poorly. Translocation of PKC-ζ from the cytosolic to the particulate membrane fraction upon exposure of monocytes to LPS provided further evidence for activation of the kinase. Preincubation of monocytes with the phosphatidylinositol 3-kinase (PI 3-kinase) inhibitors, wortmannin or LY294002, abrogated LPS-induced activation of PKC-ζ. Furthermore, activation of PKC-ζ failed to occur in U937 cells transfected with a dominant negative mutant of the p85 subunit of PI 3-kinase. PKC-ζ activity was also observed to be enhanced in vitro by the addition of phosphatidylinositol 3,4,5P3. These findings are consistent with a model in which PKC-ζ is activated downstream of PI 3-kinase in monocytes in response to LPS.
Brood parasites lay eggs matching the appearance of host clutches

Interspecific brood parasitism represents a prime example of the coevolutionary arms race where each party has evolved strategies in response to the other. Here, we investigated whether common cuckoos (Cuculus canorus) actively select nests within a host population to match the egg appearance of a particular host clutch. To achieve this goal, we quantified the degree of egg matching using the avian vision modelling approach. Randomization tests revealed that cuckoo eggs in naturally parasitized nests showed lower chromatic contrast to host eggs than those assigned randomly to other nests with egg-laying date similar to naturally parasitized clutches. Moreover, egg matching in terms of chromaticity was better in naturally parasitized nests than it would be in the nests of the nearest active non-parasitized neighbour. However, there was no indication of matching in achromatic spectral characteristics whatsoever. Thus, our results clearly indicate that cuckoos select certain host nests to increase matching of their own eggs with host clutches, but only in chromatic characteristics. Our results suggest that the ability of cuckoos to actively choose host nests based on the eggshell appearance imposes a strong selection pressure on host egg recognition.
Targeted expression of anti‐apoptotic protein p35 in oligodendrocytes reduces delayed demyelination and functional impairment after spinal cord injury

Functional impairment after spinal cord injury (SCI) is attributed to neuronal cell necrosis death and axonotmesis, with further worsening caused by the accompanying apoptosis of myelin‐forming oligodendrocytes (OLGs). However, it is unclear as to how much OLG apoptosis contributes to functional impairment. To address this issue, we used transgenic mice characterized by the targeted expression of p35, a broad‐spectrum caspase inhibitor, in OLGs using the cre/loxP system (referred to as cre/p35 transgenic mice). In this study, we examined the motor function and histopathologic changes after a contusive thoracic spinal cord injury in the cre/p35 transgenic mice. A larger number of OLGs and a lesser extent of demyelination were observed after SCI in the cre/p35 transgenic mice than in the control cre mice, which did not carry the p35 transgene. Furthermore, the motor function of the hindlimbs recovered to a significantly better degree in the cre/p35 transgenic mice than in the control cre mice. Thus, the inhibition of OLG apoptosis decreased the extent of functional impairment after SCI. These findings suggest that the inhibition of OLG apoptosis may be a potential treatment for SCI. © 2005 Wiley‐Liss, Inc.
Demonstration and characterization of dipeptide transport system activity in sheep omasal epithelium by expression of mRNA in Xenopus laevis oocytes.

Research from this laboratory has recently demonstrated that the omasal epithelium of sheep is capable of absorbing dipeptides. In order to express proteins potentially responsible for the mediated absorption of small peptides, size-fractionated poly(A)+RNA (RNA) isolated from omasal epithelial tissue of sheep (average BW 67.5 kg) were injected into defolliculated Xenopus laevis oocytes. The ability of oocytes injected with RNA or water to absorb [14C]glycyl-L-sarcosine (Gly-Sar) from media (usually pH 5.5) was compared. After 4 d (P < .02) of culture, specific RNA fractions induced an increased (P < .02) rate of Gly-Sar absorption, as compared with water-injected oocytes. The dependency of Gly-Sar uptake on the presence of a pH gradient was evaluated at pH 5.0, 5.5, 6.0, 6.5, and 7.5. Inducible uptake increased (P < .001) in the presence of increasing proton concentrations, whereas endogenous uptake of Gly-Sar decreased (P < .001). At pH 5.5, induced Gly-Sar uptake was saturable (Kt = .4 mM), but endogenous uptake was not. The specificity of Gly-Sar absorption was studied by the co-incubation of . 1 mM Gly-Sar with 5 mM levels of competing substrates (pH 5.5). Induced uptake was inhibited (P < .05) 44% by carnosine, 94% by methionylglycine, and 91% by glycylleucine, but not by glycine. Incubation of RNA with DNA oligomers that were complementary to the rabbit intestinal transporter completely inhibited (P < .05) induced Gly-Sar uptake. These results indicate that sheep omasal epithelial cells express messenger RNA that encode for proteins that are capable of H(+)-dependent dipeptide transport activity.
Apoptosis in mammalian preimplantation embryos: Regulation by survival factors

The formation of a developmentally competent mammalian blastocyst requires the transition from a unicellular state, the fertilized zygote, to a differentiated multicellular structure. In common with other developing organisms, generation of the required cell population involves the processes of cell division, differentiation and cell death, all of which can be regulated by peptide growth factors. Cell death in the preimplantation embryo occurs by apoptosis and, by analogy with other systems, may serve to eliminate unwanted cells during the critical developmental transitions that take place during this period. Cells may be eliminated because they are abnormal or possess defects, including damaged DNA or chromosomal abnormalities. At the early cleavage stages, apoptosis may be associated with activation of the embryonic genome and may contribute to the blastomere fragmentation commonly observed in human IVF embryos. The major wave of apoptosis occurs in a number of species in the inner cell mass of the blastocyst, as identified using nuclear labelling including terminal transferasemediated dUTP nick end labelling (TUNEL) and fluorescence and confocal microscopy. Apoptosis may protect the integrity and cellular composition of the inner cell mass, by eliminating damaged cells or possibly those with an inappropriate phenotype. Preimplantation embryos express genes involved in the regulation and execution of apoptosis and their cells can undergo this default pathway in the absence of exogenous survival signals. Evidence is now accumulating from several species that apoptosis in the embryo is regulated by soluble peptide growth factors acting as survival factors in an autocrine or paracrine manner. To date, these include transformaing growth factor α and members of the insulin-like growth factor family. Apoptosis may also be affected by environmental factors, including culture conditions and the composition of media. The regulation of apoptosis in the preimplantation embryo is likely to be of critical importance for both embryo viability and for later development, since the cells of the inner cell mass give rise to the fetus and carry the germ line.
Novel Function of the Transactivation Domain of a Pituitary-specific Transcription Factor, Pit-1*

Pit-1 stimulates the expression of growth hormone, prolactin, and thyrotropin β subunit genes. Consequently, abnormality of the Pit-1 gene results in combined pituitary hormone deficiency (CPHD). In this study, we analyzed the function of Pit-1 with a mutation (proline to leucine at codon 24) in the transactivation domain, P24L, which has a normal POU domain important for binding to DNA, because this mutation had been reported in a patient with CPHD. We found that codon 24 proline in the transactivation domain as well as the POU domain of Pit-1 was crucial to recruit coactivator CREB-binding protein (CBP) in the cultured cells. P24L completely lost the responsiveness to cAMP to stimulate the expression of the Pit-1-targeted genes. Furthermore, CBP and Pit-1, but not P24L, markedly enhanced the expression of the Pit-1-targeted gene to cAMP, and adenovirus E1a that binds to CBP and abrogates its function blocked the induction by cAMP of Pit-1-stimulated gene transcription in the pituitary-derived GH3 cells. These results suggest that CBP and proline at codon 24 in the transactivation domain of Pit-1 are important for the cAMP-induced activation of Pit-1-targeted genes. However, P24L maintained basal transcriptional activity, suggesting that CBP is unlikely to be an essential coactivator for Pit-1.