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971.
Tumor necrosis factor (TNF) and members of the interferon (IFN) family have been shown to independently inhibit the replication of a variety of viruses. In addition, previous reports have shown that treatment with various combinations of these antiviral cytokines induces a synergistic antiviral state that can be significantly more potent than addition of any of these cytokines alone. The mechanism of this cytokine synergy and its effects on global gene expression, however, are not well characterized. Here, we use DNA microarray analysis to demonstrate that treatment of uninfected primary human fibroblasts with TNF plus IFN-β induces a distinct synergistic state characterized by significant perturbations of several hundred genes which are coinduced by the individual cytokines alone, as well as the induction of more than 850 novel host cell genes. This synergy is mediated directly by the two ligands, not by intermediate secreted factors, and is necessary and sufficient to completely block the productive replication and spread of myxoma virus in human fibroblasts. In contrast, the replication of two other poxviruses, vaccinia virus and tanapox virus, are only partially inhibited in these cells by the synergistic antiviral state, whereas the spread of both of these viruses to neighboring cells was efficiently blocked. Taken together, our data indicate that the combination of TNF and IFN-β induces a novel synergistic antiviral state that is highly distinct from that induced by either cytokine alone. 相似文献
972.
Myotonic dystrophy 1 (MD1) is caused by a CTG expansion in the 3′-unstranslated region of the myotonic dystrophy protein kinase (DMPK) gene. MD1 patients frequently present insulin resistance and increased visceral adiposity. We examined whether DMPK deficiency is a genetic risk factor for high-fat diet-induced adiposity and insulin resistance using the DMPK knockout mouse model. We found that high-fat fed DMPK knockout mice had significantly increased body weights, hypertrophic adipocytes and whole-body insulin resistance compared with wild-type mice. This nutrient-genome interaction should be considered by physicians given the cardiometabolic risks and sedentary lifestyle associated with MD1 patients. 相似文献
973.
Nik A. B. N. Mahmood Esther Biemans-Oldehinkel Bert Poolman 《The Journal of biological chemistry》2009,284(21):14368-14376
We have previously shown that the C-terminal cystathionine β-synthase
(CBS) domains of the nucleotide-binding domains of the ABC transporter OpuA,
in conjunction with an anionic membrane surface function, act as sensor of
internal ionic strength (Iin). Here, we show that a
surface-exposed cationic region in the CBS module domain is critical for ion
sensing. The consecutive substitution of up to five cationic residues led to a
gradual decrease of the ionic strength dependence of transport. In fact, a
5-fold mutant was essentially independent of salt in the range from 0 to 250
mm KCl (or NaCl), supplemented to medium of 30 mm
potassium phosphate. Importantly, the threshold temperature for transport was
lowered by 5–7 °C and the temperature coefficient
Q10 was lowered from 8 to ∼1.5 in the 5-fold mutant,
indicating that large conformational changes are accompanying the CBS-mediated
regulation of transport. Furthermore, by replacing the anionic C-terminal tail
residues that extend the CBS module with histidines, the transport of OpuA
became pH-dependent, presumably by additional charge interactions of the
histidine residues with the membrane. The pH dependence was not observed at
high ionic strength. Altogether the analyses of the CBS mutants support the
notion that the osmotic regulation of OpuA involves a simple biophysical
switching mechanism, in which nonspecific electrostatic interactions of a
protein module with the membrane are sufficient to lock the transporter in the
inactive state.In their natural habitats microorganisms are often exposed to changes in
the concentration of solutes in the environment
(1). A sudden increase in the
medium osmolality results in loss of water from the cell, loss of turgor, a
decrease in cell volume, and an increase in intracellular osmolyte
concentration. Osmoregulatory transporters such as OpuA in Lactococcus
lactis, ProP in Escherichia coli, and BetP in
Corynebacterium glutamicum diminish the consequences of the osmotic
stress by mediating the uptake of compatible solutes upon an increase in
extracellular osmolality
(2–4).
For the ATP-binding cassette
(ABC)5 transporter
OpuA, it has been shown that the system, reconstituted in proteoliposomes, is
activated by increased concentrations of lumenal ions (increased internal
ionic strength) (2,
5,
6). This activation is
instantaneous both in vivo and in vitro and only requires
threshold levels of ionic osmolytes. Moreover, the ionic threshold for
activation is highly dependent of the ionic lipid content (charge density) of
the membrane and requires the presence of so-called cystathionine
β-synthase (CBS) domains, suggesting that the ionic signal is transduced
to the transporter via critical interactions of the protein with membrane
lipids.The ABC transporter OpuA consists of two identical nucleotide-binding
domains (NBD) fused to CBS domains and two identical substrate-binding domains
fused to transmembrane domains. The NBD-CBS and substrate-binding
domain-transmembrane domain subunits are named OpuAA and OpuABC, respectively.
Two tandem CBS domains are linked to the C-terminal end of the NBD; each
domain (CBS1 and CBS2) has a β-α-β-β-α secondary
structure (5)
(Fig. 1A). The CBS
domains are widely distributed in most if not all species of life but their
function is largely unknown. Most of the CBS domains are found as tandem
repeats but data base searches have also revealed tetra-repeat units
(5). The crystal structures of
several tandem CBS domains have been elucidated
(7–9,
32), and in a number of cases
it has been shown that two tandem CBS domains form dimeric structures with a
total of four CBS domains per structural module (hereafter referred to as CBS
module). The crystal structures of the full-length MgtE Mg2+
transporter confirm the dimeric configuration and show that the CBS domains
undergo large conformational changes upon Mg2+ binding or release
(10,
11). In general, ABC
transporters are functional as dimers, which implies that two tandem CBS
domains are present in the OpuA complex. Preliminary experiments with
disulfides engineered at the interface of two tandem CBS domains in OpuA
suggest that large structural rearrangements (association-dissociation of the
interfaces) play a determining role in the ionic strength-regulated transport.
Finally, a subset of CBS-containing proteins has a C-terminal extension, which
in OpuA is highly anionic (sequence: ADIPDEDEVEEIEKEEENK) and modulates the
ion sensing activity (6).Open in a separate windowFIGURE 1.Domain structure of CBS module of OpuA. A, sequence of
tandem CBS domains. The predicted secondary structure is indicated
above the sequence. The residues modified in this study are
underlined. The amino acid sequence end-points of OpuAΔ61 and
OpuAΔ119 are indicated by vertical arrows. B, homology
model of tandem CBS domain of OpuA. The CBS domains were individually modeled
on the crystal structure of the tandem CBS protein Ta0289 from T.
acidophilum (PDB entry 1PVM), using Phyre. Ta0289 was used for the
initial modeling, because its primary sequence was more similar to the CBS
domains of OpuA than those of the other crystallized CBS proteins. The
individual domain models were then assembled with reference to the atomic
coordinates of the tandem CBS domains of IMPDH from Streptococcus
pyogenes (PDB entry 1ZFJ) to form the tandem CBS pair, using PyMOL
(DeLano). The positions of the (substituted) cationic residues are
indicated.In this study, we have engineered the surface-exposed cationic residues of
the CBS module and the C-terminal anionic tail of OpuA
(Fig. 1B). The ionic
strength and lipid dependence of the OpuA mutants were determined in
vivo and in vitro. We show that substitution of five cationic
residues for neutral amino acids is sufficient to inactivate the ionic
strength sensor and convert OpuA into a constitutively active transporter.
Moreover, by substituting six anionic plus four neutral residues of the
C-terminal anionic tail for histidines, the transport reaction becomes
strongly pH-dependent. 相似文献
974.
Christine Decaestecker Xavier Moles Lopez Nicky D'Haene Isabelle Roland Saad Guendouz Christophe Duponchelle Alix Berton Olivier Debeir Isabelle Salmon 《Proteomics》2009,9(19):4478-4494
Antibody‐based proteomics applied to tissue microarray (TMA) technology provides a very efficient means of visualizing and locating antigen expression in large collections of normal and pathological tissue samples. To characterize antigen expression on TMAs, the use of image analysis methods avoids the effects of human subjectivity evidenced in manual microscopical analysis. Thus, these methods have the potential to significantly enhance both precision and reproducibility. Although some commercial systems include tools for the quantitative evaluation of immunohistochemistry‐stained images, there exists no clear agreement on best practices to allow for correct and reproducible quantification results. Our study focuses on practical aspects regarding (i) image acquisition (ii) segmentation of staining and counterstaining areas and (iii) extraction of quantitative features. We illustrate our findings using a commercial system to quantify different immunohistochemistry markers targeting proteins with different expression patterns (cytoplasmic, nuclear or membranous) in colon cancer or brain tumor TMAs. Our investigations led us to identify several steps that we consider essential for standardizing computer‐assisted immunostaining quantification experiments. In addition, we propose a data normalization process based on reference materials to be able to compare measurements between studies involving different TMAs. In conclusion, we recommend certain critical prerequisites that commercial or in‐house systems should satisfy in order to permit valid immunostaining quantification. 相似文献
975.
Aiqing Li Joshua B. Benoit Giancarlo Lopez‐Martinez Michael A. Elnitsky Richard E. Lee Jr David L. Denlinger 《Proteomics》2009,9(10):2788-2798
Desiccation presents a major challenge for the Antarctic midge, Belgica antarctica. In this study, we use proteomic profiling to evaluate protein changes in the larvae elicited by dehydration and rehydration. Larvae were desiccated at 75% relative humidity (RH) for 12 h to achieve a body water loss of 35%, approximately half of the water that can be lost before the larvae succumb to dehydration. To evaluate the rehydration response, larvae were first desiccated, then rehydrated for 6 h at 100% RH and then in water for 6 h. Controls were held continuously at 100% RH. Protein analysis was performed using 2‐DE and nanoscale capillary LC/MS/MS. Twenty‐four identified proteins changed in abundance in response to desiccation: 16 were more abundant and 8 were less abundant; 84% of these proteins were contractile or cytoskeletal proteins. Thirteen rehydration‐regulated proteins were identified: 8 were more abundant and 5 were less abundant, and 69% of these proteins were also contractile or cytoskeletal proteins. Additional proteins responsive to desiccation and rehydration were involved in functions including stress responses, energy metabolism, protein synthesis, glucogenesis and membrane transport. We conclude that the major protein responses elicited by both desiccation and rehydration are linked to body contraction and cytoskeleton rearrangements. 相似文献
976.
Integration of tomato reproductive developmental landmarks and expression profiles, and the effect of SUN on fruit shape 总被引:1,自引:0,他引:1
Han Xiao Cheryll Radovich Nicholas Welty Jason Hsu Dongmei Li Tea Meulia Esther van der Knaap 《BMC plant biology》2009,9(1):49-21
Background
Universally accepted landmark stages are necessary to highlight key events in plant reproductive development and to facilitate comparisons among species. Domestication and selection of tomato resulted in many varieties that differ in fruit shape and size. This diversity is useful to unravel underlying molecular and developmental mechanisms that control organ morphology and patterning. The tomato fruit shape gene SUN controls fruit elongation. The most dramatic effect of SUN on fruit shape occurs after pollination and fertilization although a detailed investigation into the timing of the fruit shape change as well as gene expression profiles during critical developmental stages has not been conducted. 相似文献977.
Antonio Cabrera Alex Kozik Werner Howad Pere Arus Amy F Iezzoni Esther van der Knaap 《BMC genomics》2009,10(1):562
Background
Detailed comparative genome analyses within the economically important Rosaceae family have not been conducted. This is largely due to the lack of conserved gene-based molecular markers that are transferable among the important crop genera within the family [e.g. Malus (apple), Fragaria (strawberry), and Prunus (peach, cherry, apricot and almond)]. The lack of molecular markers and comparative whole genome sequence analysis for this family severely hampers crop improvement efforts as well as QTL confirmation and validation studies.Results
We identified a set of 3,818 rosaceaous unigenes comprised of two or more ESTs that correspond to single copy Arabidopsis genes. From this Rosaceae Conserved Orthologous Set (RosCOS), 1039 were selected from which 857 were used for the development of intron-flanking primers and allele amplification. This led to successful amplification and subsequent mapping of 613 RosCOS onto the Prunus TxE reference map resulting in a genome-wide coverage of 0.67 to 1.06 gene-based markers per cM per linkage group. Furthermore, the RosCOS primers showed amplification success rates from 23 to 100% across the family indicating that a substantial part of the RosCOS primers can be directly employed in other less studied rosaceaous crops. Comparisons of the genetic map positions of the RosCOS with the physical locations of the orthologs in the Populus trichocarpa genome identified regions of colinearity between the genomes of Prunus-Rosaceae and Populus-Salicaceae.Conclusion
Conserved orthologous genes are extremely useful for the analysis of genome evolution among closely and distantly related species. The results presented in this study demonstrate the considerable potential of the mapped Prunus RosCOS for genome-wide marker employment and comparative whole genome studies within the Rosaceae family. Moreover, these markers will also function as useful anchor points for the genome sequencing efforts currently ongoing in this family as well as for comparative QTL analyses.978.
979.
980.
Marco P. Boks Eske M. Derks Daniel J. Weisenberger Erik Strengman Esther Janson Iris E. Sommer René S. Kahn Roel A. Ophoff 《PloS one》2009,4(8)
Cytosine-5 methylation within CpG dinucleotides is a potentially important mechanism of epigenetic influence on human traits and disease. In addition to influences of age and gender, genetic control of DNA methylation levels has recently been described. We used whole blood genomic DNA in a twin set (23 MZ twin-pairs and 23 DZ twin-pairs, N = 92) as well as healthy controls (N = 96) to investigate heritability and relationship with age and gender of selected DNA methylation profiles using readily commercially available GoldenGate bead array technology. Despite the inability to detect meaningful methylation differences in the majority of CpG loci due to tissue type and locus selection issues, we found replicable significant associations of DNA methylation with age and gender. We identified associations of genetically heritable single nucleotide polymorphisms with large differences in DNA methylation levels near the polymorphism (cis effects) as well as associations with much smaller differences in DNA methylation levels elsewhere in the human genome (trans effects). Our results demonstrate the feasibility of array-based approaches in studies of DNA methylation and highlight the vast differences between individual loci. The identification of CpG loci of which DNA methylation levels are under genetic control or are related to age or gender will facilitate further studies into the role of DNA methylation and disease. 相似文献