共查询到20条相似文献,搜索用时 15 毫秒
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Akifumi Sugiyama Shoju Fukuda Kojiro Takanashi Miki Yoshioka Hirofumi Yoshioka Yoshihiro Narusaka Mari Narusaka Mikiko Kojima Hitoshi Sakakibara Nobukazu Shitan Shusei Sato Satoshi Tabata Masayoshi Kawaguchi Kazufumi Yazaki 《PloS one》2015,10(9)
LjABCG1, a full-size ABCG subfamily of ATP-binding cassette proteins of a model legume, Lotus japonicus, was reported as a gene highly expressed during the early stages of nodulation, but have not been characterized in detail. In this study we showed that the induction of LjABCG1 expression was remarkable by methyl jasmonate treatment, and reporter gene experiments indicated that LjABCG1 was strongly expressed in the nodule parenchyma and cell layers adjacent to the root vascular tissue toward the nodule. LjABCG1 was suggested to be localized at the plasma membrane based on the fractionation of microsomal membranes as well as separation via aqueous two-phase partitioning. The physiological functions of LjABCG1 in symbiosis and pathogenesis were analyzed in homologous and heterologous systems. LjABCG1 knock-down L. japonicus plants did not show clear phenotypic differences in nodule formation, and not in defense against Pseudomonas syringae, either. In contrast, when LjABCG1 was expressed in the Arabidopsis pdr8-1 mutant, the penetration frequency of Phytophthora infestans, a potato late blight pathogen, was significantly reduced in LjABCG1/pdr8-1 than in pdr8-1 plants. This finding indicated that LjABCG1, at least partially, complemented the phenotype of pdr8 in Arabidopsis, suggesting the multiple roles of this protein in plant-microbe interactions. 相似文献
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Myriam Charpentier Rolf Bredemeier Gerhard Wanner Naoya Takeda Enrico Schleiff Martin Parniske 《The Plant cell》2008,20(12):3467-3479
The mechanism underlying perinuclear calcium spiking induced during legume root endosymbioses is largely unknown. Lotus japonicus symbiosis-defective castor and pollux mutants are impaired in perinuclear calcium spiking. Homology modeling suggested that the related proteins CASTOR and POLLUX might be ion channels. Here, we show that CASTOR and POLLUX form two independent homocomplexes in planta. CASTOR reconstituted in planar lipid bilayers exhibited ion channel activity, and the channel characteristics were altered in a symbiosis-defective mutant carrying an amino acid replacement close to the selectivity filter. Permeability ratio determination and competition experiments reveled a weak preference of CASTOR for cations such as potassium over anions. POLLUX has an identical selectivity filter region and complemented a potassium transport–deficient yeast mutant, suggesting that POLLUX is also a potassium-permeable channel. Immunogold labeling localized the endogenous CASTOR protein to the nuclear envelope of Lotus root cells. Our data are consistent with a role of CASTOR and POLLUX in modulating the nuclear envelope membrane potential. They could either trigger the opening of calcium release channels or compensate the charge release during the calcium efflux as counter ion channels. 相似文献
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David Dauvillée Christophe Colleoni Eudean Shaw Gregory Mouille Christophe D''Hulst Matthew Morell Michael S. Samuel Brigitte Bouchet Daniel J. Gallant Anthony Sinskey Steven Ball 《Plant physiology》1999,119(1):321-330
We isolated two muskmelon (Cucumis melo) cDNA homologs of the Arabidopsis ethylene receptor genes ETR1 and ERS1 and designated them Cm-ETR1 (C. melo ETR1; accession no. AF054806) and Cm-ERS1 (C. melo ERS1; accession no. AF037368), respectively. Northern analysis revealed that the level of Cm-ERS1 mRNA in the pericarp increased in parallel with the increase in fruit size and then markedly decreased at the end of enlargement. In fully enlarged fruit the level of Cm-ERS1 mRNA was low in all tissues, whereas that of Cm-ETR1 mRNA was very high in the seeds and placenta. During ripening Cm-ERS1 mRNA increased slightly in the pericarp of fruit before the marked increase of Cm-ETR1 mRNA paralleled climacteric ethylene production. These results indicate that both Cm-ETR1 and Cm-ERS1 play specific roles not only in ripening but also in the early development of melon fruit and that they have distinct roles in particular fruit tissues at particular developmental stages. 相似文献
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Primary roots of tomato, Lycopersicon esculentum cv. Marglobe, were cultured aseptically on agar containing a standard nutrient formulation with or without kinetin. When secondary roots developed, cultures were inoculated with the root-knot nematode, Meloidogyne incognita. Following inoculation, the cultures were divided into two groups which were incubated either in total darkness or in 16-h light-8-h dark cycles. At 24 h, 1, 2, 3, and 4 wk after incubation, roots from all cultures were processed for transmission electron microscopy. Fine structural observation of the parenchyma tissue in galls from the inoculated cultures indicated that starch containing plastids or amyloplasts, which are usually present and remain undifferentiated in these root cells, developed into chloroplasts. These chloroplasts contained a membrane system indistinguishable from those found in leaves of intact plants. Although plastid development was not affected when uninoculated cultures were incubated in the light, differentiation of the amyloplast was induced when roots were cultured on the medium containing kinetin. These results suggest that plastid differentiation in the inoculated tissue may be influenced by an accumulation of kinetin in the gall, which is induced by the nematode and serves as the nutrient sink for its feeding. 相似文献
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Linlin Xing Hui Yu Jingjing Qi Pan Jiang Bingqing Sun Junsheng Cui Changcan Ou Weishan Chang Qinghai Hu 《PloS one》2015,10(6)
To investigate the genetic basis of erythromycin resistance in Riemerella anatipestifer, the MIC to erythromycin of 79 R. anatipestifer isolates from China and one typed strain, ATCC11845, were evaluated. The results showed that 43 of 80 (53.8%) of the tested R. anatipestifer strains showed resistance to erythromycin, and 30 of 43 erythromycin-resistant R. anatipestifer strains carried ermF or ermFU with an MIC in the range of 32–2048 μg/ml, while the other 13 strains carrying the ereD gene exhibited an MIC of 4–16 μg/ml. Of 30 ermF + R. anatipestifer strains, 27 (90.0%) carried the ermFU gene which may have been derived from the CTnDOT-like element, while three other strains carried ermF from transposon Tn4351. Moreover, sequence analysis revealed that ermF, ermFU, and ereD were located within the multiresistance region of the R. anatipestifer genome. 相似文献
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Jeffrey M. Boyd Jamie L. Sondelski Diana M. Downs 《The Journal of biological chemistry》2009,284(1):110-118
The ApbC protein has been shown previously to bind and rapidly transfer
iron-sulfur ([Fe-S]) clusters to an apoprotein (Boyd, J. M., Pierik, A. J.,
Netz, D. J., Lill, R., and Downs, D. M. (2008) Biochemistry 47,
8195–8202. This study utilized both in vivo and in
vitro assays to examine the function of variant ApbC proteins. The in
vivo assays assessed the ability of ApbC proteins to function in pathways
with low and high demand for [Fe-S] cluster proteins. Variant ApbC proteins
were purified and assayed for the ability to hydrolyze ATP, bind [Fe-S]
cluster, and transfer [Fe-S] cluster. This study details the first kinetic
analysis of ATP hydrolysis for a member of the ParA subfamily of
“deviant” Walker A proteins. Moreover, this study details the
first functional analysis of mutant variants of the ever expanding family of
ApbC/Nbp35 [Fe-S] cluster biosynthetic proteins. The results herein show that
ApbC protein needs ATPase activity and the ability to bind and rapidly
transfer [Fe-S] clusters for in vivo function.Proteins containing iron-sulfur ([Fe-S]) clusters are employed in a wide
array of metabolic functions (reviewed in Ref.
1). Research addressing the
biosynthesis of the iron-molybdenum cofactor of nitrogenase in Azotobacter
vinelandii led to the discovery of an operon
(iscAnifnifUSVcysE1) involved in the
biosynthesis of [Fe-S] clusters (reviewed in Ref.
2). Subsequent experiments led
to the finding of two more systems involved in the de novo
biosynthesis of [Fe-S] clusters, the isc and the suf systems
(3,
4). Like Escherichia
coli, the genome of Salmonella enterica serovar Typhimurium
encodes for the isc and suf [Fe-S] cluster biosynthesis
machinery.Recent studies have identified a number of additional or
non-isc/-suf-encoded proteins that are involved in bacterial
[Fe-S] cluster biosynthesis and repair. Examples include the following: CyaY,
an iron-binding protein believed to be involved in iron trafficking and iron
delivery
(5–7);
YggX, an Fe2+-binding protein that protects the cell from oxidative
stress (8,
9); ErpA, an alternate A-type
[Fe-S] cluster scaffolding protein
(10); NfuA, a proposed
intermediate [Fe-S] delivery protein
(11–13);
YtfE, a protein proposed to be involved in [Fe-S] cluster repair
(14,
15); and CsdA-CsdE, an
alternative cysteine desulferase
(16).Analysis of the metabolic network anchored to thiamine biosynthesis in
S. enterica identified lesions in three non-isc or
-suf loci that compromise Fe-S metabolism as follows: apbC,
apbE, and rseC
(17–21).
This metabolic system was subsequently used to dissect a role for
cyaY and gshA in [Fe-S] cluster metabolism
(6,
22,
23). Of these, the
apbC (mrp in E. coli) locus was identified as the
predominant site of lesions that altered thiamine synthesis by disrupting
[Fe-S] cluster metabolism (17,
18).ApbC is a member of the ParA subfamily of proteins that have a wide array
of functions, including electron transfer
(24), initiation of cell
division (25), and DNA
segregation (26,
27). Importantly, ATP
hydrolysis is required for function of all well characterized members of this
subfamily, and all members contain a “deviant” Walker A motif,
which contains two lysine residues instead of one (GKXXXGK(S/T))
(28). ApbC has been shown to
hydrolyze ATP (17).Recently, five proteins with a high degree of identity to ApbC have been
shown to be involved in [Fe-S] cluster metabolism in eukaryotes. The sequence
alignments of the central portion of these proteins and bacterial ApbC are
shown in Fig. 1. HCF101 was
demonstrated to be involved in chloroplast [Fe-S] cluster metabolism
(29,
30). The CFD1, Npb35, and
huNbp35 (formally Nubp1) proteins were demonstrated to be involved in
cytoplasmic [Fe-S] cluster metabolism
(31,
32). Ind1 was demonstrated to
be involved in the maturation of [Fe-S] clusters in the mitochondrial enzyme
NADH:ubiquinone oxidoreductase
(33). There is currently no
report of any of these proteins hydrolyzing ATP.Open in a separate windowFIGURE 1.Protein sequence alignments of members of the ApbC/Nbp35 subfamily of
ParA family of proteins. Protein alignments were assembled using the
Clustal_W method in the Lasergene® software and show only the central
portion of the proteins, which have the highest sequence conservation. The
three boxed areas highlight the Walker A box, conserved Ser residue,
and CXXC motif. Proteins listed are as follows: ApbC (S.
enterica serovar Typhimurium LT2), CFD1 (S. cerevisiae), Nbp35
(S. cerevisiae), HCF101 (Arabidopsis thaliana), huNpb35
(formally Nubp1) (Homo sapiens), and Ind1 (Candida
albicans).Biochemical analysis of ApbC indicated that it could bind and transfer
[Fe-S] clusters to Saccharomyces cerevisiae apo-isopropylmalate
isomerase (34). Additional
genetic studies indicated that ApbC has a degree of functional redundancy with
IscU, a known [Fe-S] cluster scaffolding protein
(35,
36).In this study we investigate the correlation between the biochemical
properties of ApbC (i.e. ATPase activity, [Fe-S] cluster binding, and
[Fe-S] cluster transfer rates) and the in vivo function of this
protein. This is the first detailed kinetic analysis of ATP hydrolysis for a
member of the ParA subfamily of deviant Walker A proteins and the first
functional analysis of a member of the ever expanding family of ApbC/Nbp35
proteins. Data presented indicate that noncomplementing variants have distinct
biochemical properties that place them in three distinct classes. 相似文献
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Line Lapointe 《Plant physiology》1998,117(1):183-188
Leaves are the main source of carbon for fruit maturation in most species. However, in plants seeing contrasting light conditions such as some spring plants, carbon fixed during the spring could be used to support fruit development in the summer, when photosynthetic rates are low. We monitored carbohydrate content in the rhizome (a perennating organ) and the aboveground stem of trillium (Trillium erectum) over the entire growing season (May–November). At the beginning of the fruiting stage, stems carrying a developing fruit were harvested, their leaves were removed, and the leafless stems were maintained in aqueous solution under controlled conditions up to full fruit maturation. These experiments showed that stem carbohydrate content was sufficient to support fruit development in the absence of leaves and rhizome. This is the first reported case, to our knowledge, of complete fruit development sustained only by a temporary carbohydrate reservoir. This carbohydrate accumulation in the stem during the spring enables the plant to make better use of the high irradiances occurring at that time. Many other species might establish short-term carbohydrate reservoirs in response to seasonal changes in growing conditions. 相似文献
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Yang Wang Dan Li Roza Nurieva Justin Yang Mehmet Sen Roberto Carre?o Sijie Lu Bradley W. McIntyre Jeffrey J. Molldrem Glen B. Legge Qing Ma 《The Journal of biological chemistry》2009,284(19):12645-12653
The activation of LFA-1 (lymphocyte function-associated antigen) is a
critical event for T cell co-stimulation. The mechanism of LFA-1 activation
involves both affinity and avidity regulation, but the role of each in T cell
activation remains unclear. We have identified antibodies that recognize and
block different affinity states of the mouse LFA-1 I-domain. Monoclonal
antibody 2D7 preferentially binds to the low affinity conformation, and this
specific binding is abolished when LFA-1 is locked in the high affinity
conformation. In contrast, M17/4 can bind both the locked high and low
affinity forms of LFA-1. Although both 2D7 and M17/4 are blocking antibodies,
2D7 is significantly less potent than M17/4 in blocking LFA-1-mediated
adhesion; thus, blocking high affinity LFA-1 is critical for preventing
LFA-1-mediated adhesion. Using these reagents, we investigated whether LFA-1
affinity regulation affects T cell activation. We found that blocking high
affinity LFA-1 prevents interleukin-2 production and T cell proliferation,
demonstrated by TCR cross-linking and antigen-specific stimulation.
Furthermore, there is a differential requirement of high affinity LFA-1 in the
activation of CD4+ and CD8+ T cells. Although
CD4+ T cell activation depends on both high and low affinity LFA-1,
only high affinity LFA-1 provides co-stimulation for CD8+ T cell
activation. Together, our data demonstrated that the I-domain of LFA-1 changes
to the high affinity state in primary T cells, and high affinity LFA-1 is
critical for facilitating T cell activation. This implicates LFA-1 activation
as a novel regulatory mechanism for the modulation of T cell activation and
proliferation.LFA-1 (lymphocyte function-associated antigen), an integrin family member,
is important in regulating leukocyte adhesion and T cell activation
(1,
2). LFA-1 consists of the
αL (CD11a) and β2 (CD18) heterodimer. The
ligands for LFA-1, including intercellular adhesion molecule
ICAM3-1, ICAM-2, and
ICAM-3, are expressed on antigen-presenting cells (APCs), endothelial cells,
and lymphocytes (1). Mice that
are deficient in LFA-1 have defects in leukocyte adhesion, lymphocyte
proliferation, and tumor rejection
(3–5).
Blocking LFA-1 with antibodies can prevent inflammation, autoimmunity, organ
graft rejection, and graft versus host disease in human and murine
models
(6–10).LFA-1 is constitutively expressed on the surface of leukocytes in an
inactive state. Activation of LFA-1 is mediated by inside-out signals from the
cytoplasm (1,
11). Subsequently, activated
LFA-1 binds to the ligands and transduces outside-in signals back into the
cytoplasm that result in cell adhesion and activation
(12,
13). The activation of LFA-1
is a critical event in the formation of the immunological synapse, which is
important for T cell activation
(2,
14,
15). The active state of LFA-1
is regulated by chemokines and the T cell receptor (TCR) through Rap1
signaling (16). LFA-1 ligation
lowers the activation threshold and affects polarization in CD4+ T
cells (17). Moreover,
productive LFA-1 engagement facilitates efficient activation of cytotoxic T
lymphocytes and initiates a distinct signal essential for the effector
function
(18–20).
Thus, LFA-1 activation is essential for the optimal activation of T cells.The mechanism of LFA-1 activation involves both affinity (conformational
changes within the molecule) and avidity (receptor clustering) regulation
(21–23).
The I-domain of the LFA-1 αL subunit is the primary
ligand-binding site and has been proposed to change conformation, leading to
an increased affinity for ligands
(24–26).
The structural basis of the conformational changes in the I-domain of LFA-1
has been extensively characterized
(27). Previously, we have
demonstrated that the conformation of the LFA-1 I-domain changes from the low
affinity to the high affinity state upon activation. By introducing disulfide
bonds into the I-domain, LFA-1 can be locked in either the closed or open
conformation, which represents the “low affinity” or “high
affinity” state, respectively
(28,
29). In addition, we
identified antibodies that are sensitive to the affinity changes in the
I-domain of human LFA-1 and showed that the activation-dependent epitopes are
exposed upon activation (30).
This study supports the presence of the high affinity conformation upon LFA-1
activation in cell lines. It has been demonstrated recently that therapeutic
antagonists, such as statins, inhibit LFA-1 activation and immune responses by
locking LFA-1 in the low affinity state
(31–34).
Furthermore, high affinity LFA-1 has been shown to be important for mediating
the adhesion of human T cells
(35,
36). Thus, the affinity
regulation is a critical step in LFA-1 activation.LFA-1 is a molecule of great importance in the immune system, and its
activation state influences the outcome of T cell activation. Our previous
data using the activating LFA-1 I-domain-specific antibody MEM83 indicate that
avidity and affinity of the integrin can be coupled during activation
(37). However, whether
affinity or avidity regulation of LFA-1 contributes to T cell activation
remains controversial (23,
38,
39). Despite the recent
progress suggesting that conformational changes represent a key step in the
activation of LFA-1, there are considerable gaps to be filled. When LFA-1 is
activated, the subsequent outside-in signaling contributes to T cell
activation via immunological synapse and LFA-1-dependent signaling. It is
critical to determine whether high affinity LFA-1 participates in the
outside-in signaling and affects the cellular activation of T cells.
Nevertheless, the rapid and dynamic process of LFA-1 activation has hampered
further understanding of the role of high affinity LFA-1 in primary T cell
activation. The affinity of LFA-1 for ICAM-1 increases up to 10,000-fold
within seconds and involves multiple reversible steps
(23). In addition, the
activation of LFA-1 regulates both adhesion and activation of T cells, two
separate yet closely associated cellular functions. When LFA-1 is
constitutively expressed in the active state in mice, immune responses are
broadly impaired rather than hyperactivated, suggesting the complexity of
affinity regulation (40).
Therefore, it is difficult to dissect the mechanisms by which high affinity
LFA-1 regulates stepwise activation of T cells in the whole animal system.In the present study, we identified antibodies recognizing and blocking
different affinity states of mouse LFA-1. These reagents allowed us to
determine the role of affinity regulation in T cell activation. We found that
blocking high affinity LFA-1 inhibited IL-2 production and proliferation in T
cells. Furthermore, there is a differential requirement of high affinity LFA-1
in antigen-specific activation of CD4+ and CD8+ T cells.
The activation of CD4+ T cells depends on both high and low
affinity LFA-1. For CD8+ T cell activation, only high affinity
LFA-1 provides co-stimulation. Thus, affinity regulation of LFA-1 is critical
for the activation and proliferation of naive T cells. 相似文献
16.
Hematopoietic stem cells (HSC) must engage in a life-long balance between self-renewal and differentiation to sustain hematopoiesis. The highly conserved PIWI protein family regulates proliferative states of stem cells and their progeny in diverse organisms. A Human piwi gene (for clarity, the non-italicized “piwi” refers to the gene subfamily), HIWI (PIWIL1), is expressed in CD34+ stem/progenitor cells and transient expression of HIWI in a human leukemia cell line drastically reduces cell proliferation, implying the potential function of these proteins in hematopoiesis. Here, we report that one of the three piwi genes in mice, Miwi2 (Piwil4), is expressed in primitive hematopoetic cell types within the bone marrow. Mice with a global deletion of all three piwi genes, Miwi, Mili, and Miwi2, are able to maintain long-term hematopoiesis with no observable effect on the homeostatic HSC compartment in adult mice. The PIWI-deficient hematopoetic cells are capable of normal lineage reconstitution after competitive transplantation. We further show that the three piwi genes are dispensable during hematopoietic recovery after myeloablative stress by 5-FU. Collectively, our data suggest that the function of the piwi gene subfamily is not required for normal adult hematopoiesis. 相似文献
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