bluuebunny/crossref_metadata_2025 huggingface dataset:
Magnetic field effect on premartensitic transition in Ni–Mn–Ga alloys
Authors: Barandiaran J. M. , Chernenko V. A. , Lazpita P. , Gutiérrez J. , Orue I. , Feuchtwanger J. , Besseghini S.
Published: February 2009

Extensive measurements of the temperature dependencies of resistivity under constant magnetic fields made it possible to clarify the premartensitic transition sensitivity to applied magnetic field up to 14 T in polycrystalline and monocrystalline Ni–Mn–Ga alloys with compositions close to the stoichiometric one. A low-field minimum on the transition temperature versus magnetic field curves is found in both alloys. This minimum is attributed to the magnetic anisotropy contribution to the Clausius–Clapeyron relationship, which describes the premartensitic transition. This contribution is confirmed by magnetization loops measurements.
Low-order distributed feedback optofluidic dye laser with reduced threshold
Authors: Song Wuzhou , Vasdekis Andreas E. , Li Zhenyu , Psaltis Demetri
Published: February 2009

We report the demonstration of low order distributed feedback (DFB) optofluidic dye lasers with reduced threshold. The laser chips were realized in polydimethylsiloxane using replica molding with two masters. A comparison between first, second, and third order DFB dye lasers was performed, while the second order DFB dye laser exhibited the lowest pump threshold of 78 nJ/pulse. Compared to previous reports on higher order Bragg grating structures, the pump threshold in this work is approximately 30-fold lower than the state of the art due to the reduction in the cavity losses and the more efficient pumping configuration.
Evaluation of GaN substrates grown in supercritical basic ammonia
Authors: Saito Makoto , Yamada Hisashi , Iso Kenji , Sato Hitoshi , Hirasawa Hirohiko , Kamber Derrick S. , Hashimoto Tadao , DenBaars Steven P. , Speck James S. , Nakamura Shuji
Published: February 2009

GaN crystals grown by the basic ammonothermal method were investigated for their use as substrates for device regrowth. X-ray diffraction analysis indicated that the substrates contained multiple grains while secondary ion mass spectroscopy (SIMS) revealed a high concentration of hydrogen, oxygen, and sodium. Despite these drawbacks, the emission from the light emitting diode structures grown by metal organic chemical vapor deposition on both the c-plane and m-plane epitaxial wafers was demonstrated. The SIMS depth profiles showed that the diffusion of the alkali metal from the substrate into the epitaxial film was small, especially in the m-direction.
Different patterns of herpes simplex virus type 1 infection among college students from Cuernavaca, Mexico
Authors: Arriaga-Demeza R. Carlos , Conde-Glez Carlos J. , Uribe-Salas Felipe J. , Eguiza-Fano Sergio , García-Cisneros Santa , Sánchez-Alemán Miguel A.
Published: February 2009

The goal of this study was to determine herpes simplex virus type 1 (HSV-1) seroprevalence among college students from Morelos State University, Mexico; and to correlate their demographic and sexual behaviour characteristics with HSV-1 infection. Demographic characteristics were associated with HSV-1 among female students. Among male students, sexual behaviour characteristics were associated with HSV-1 seroprevalence. Male students who practised oral sex and did not always use a condom presented higher rates of HSV-1 infection (76.9%) than students who did not have oral sex (52.2%). A different pattern of HSV-1 infection depends on gender among Mexican students analysed.
Distinct functions for the catalytic and hemopexin domains of a
Drosophila
matrix metalloproteinase
Authors: Glasheen Bernadette M. , Kabra Aashish T. , Page-McCaw Andrea
Published: February 2009

Human matrix metalloproteinases (MMPs) are believed to contribute to tumor progression. Therapies based on inhibiting the catalytic domain of MMPs have been unsuccessful, but these studies raise the question of whether other MMP domains might be appropriate targets. The genetic dissection of domain function has been stymied in mouse because there are 24 related and partially redundant MMP genes in the mouse genome. Here, we present a genetic dissection of the functions of the hemopexin and catalytic domains of a canonical MMP in
Drosophila melanogaster
, an organism with only 2 MMPs that function nonredundantly. We compare the phenotypes of
Mmp1
null alleles with alleles that have specific hemopexin domain lesions, and we also examine phenotypes of dominant-negative mutants. We find that, although the catalytic domain appears to be required for all MMP functions including extracellular matrix remodeling of the tracheal system, the hemopexin domain is required specifically for tissue invasion events later in metamorphosis but not for tracheal remodeling. Thus, we find that this MMP hemopexin domain has an apparent specialization for tissue invasion events, a finding with potential implications for inhibitor therapies.
Sialylated multivalent antigens engage CD22in transand inhibit B cell activation
Authors: Courtney Adam H. , Puffer Erik B. , Pontrello Jason K. , Yang Zhi-Qiang , Kiessling Laura L.
Published: February 2009

CD22 is an inhibitory coreceptor on the surface of B cells that attenuates B cell antigen receptor (BCR) signaling and, therefore, B cell activation. Elucidating the molecular mechanisms underlying the inhibitory activity of CD22 is complicated by the ubiquity of CD22 ligands. Although antigens can display CD22 ligands, the receptor is known to bind to sialylated glycoproteins on the cell surface. The propinquity of CD22 and cell-surface glycoprotein ligands has led to the conclusion that the inhibitory properties of the receptor are due tocisinteractions. Here, we examine the functional consequences oftransinteractions by employing sialylated multivalent antigens that can engage both CD22 and the BCR. Exposure of B cells to sialylated antigens results in the inhibition of key steps in BCR signaling. These results reveal that antigens bearing CD22 ligands are powerful suppressors of B cell activation. The ability of sialylated antigens to inhibit BCR signaling throughtransCD22 interactions reveals a previously unrecognized role for the Siglec-family of receptors as modulators of immune signaling.
p53 represses c-Myc through induction of the tumor suppressor
miR-145
Authors: Sachdeva Mohit , Zhu Shoumin , Wu Fangting , Wu Hailong , Walia Vijay , Kumar Sumit , Elble Randolph , Watabe Kounosuke , Mo Yin-Yuan
Published: February 2009

The tumor suppressor p53 negatively regulates a number of genes, including the proto-oncogene c-Myc, in addition to activating many other genes. One mechanism of the p53-mediated c-Myc repression may involve transcriptional regulation. However, it is not clear whether microRNAs (miRNAs) play a role in the p53-mediated posttranscriptional regulation of c-Myc. In this study, we show that a putative tumor suppressor,
miR-145
, is expressed through the phosphoinositide-3 kinase (PI-3K)/Akt and p53 pathways. Importantly, p53 transcriptionally induces the expression of
miR-145
by interacting with a potential p53 response element (p53RE) in the
miR-145
promoter. We further show that c-Myc is a direct target for
miR-145
. Although
miR-145
silences the expression of c-Myc, anti-
miR-145
enhances its expression. This specific silencing of c-Myc by
miR-145
accounts at least in part for the
miR-145
-mediated inhibition of tumor cell growth both in vitro and in vivo. Finally, the blockade of
miR-145
by anti-
miR-145
is able to reverse the p53-mediated c-Myc repression. Together, these results define the role of
miR-145
in the posttranscriptional regulation of c-Myc by p53 and suggest that, as a new member of the p53 regulatory network,
miR-145
provides a direct link between p53 and c-Myc in this gene regulatory network.
Anti-laminin gamma-1 pemphigoid
Authors: Dainichi Teruki , Kurono Sadamu , Ohyama Bungo , Ishii Norito , Sanzen Noriko , Hayashi Maria , Shimono Chisei , Taniguchi Yukimasa , Koga Hiroshi , Karashima Tadashi , Yasumoto Shinichiro , Zillikens Detlef , Sekiguchi Kiyotoshi , Hashimoto Takashi
Published: February 2009

Anti-p200 pemphigoid has been characterized by autoantibodies to an unidentified 200-kDa protein (p200) of the dermal−epidermal junction. The objective of this study was to identify p200. We performed 2D gel electrophoresis of dermal extracts and immunoblotting with patients' sera, followed by MS analysis of a unique protein band. The protein band corresponded to laminin γ1. Anti-laminin γ1 mAb reacted with the anti-p200 immunoprecipitates by immunoblotting. Sera from 32 patients with anti-p200 pemphigoid showed 90% reactivity to the recombinant products of laminin γ1. None of the healthy control sera reacted with laminin γ1. By immunoblotting, reactivity of a patient's serum with p200 was competitively inhibited by adding anti-laminin γ1
C
-terminus mAb. Purified anti-p200 IgG also inhibited the reactivity of this mAb to dermal laminin γ1. Most laminin γ1-positive sera showed reactivity with recombinant laminin γ1
C
-terminal E8 fragment. Reactivity of patients' sera and purified IgG to dermal laminin γ1 was higher than reactivity to blood vessel laminin γ1 under reducing conditions. These results suggest that laminin γ1 is the autoantigen for patients with anti-p200 pemphigoid. The autoantibodies may specifically recognize dermal laminin γ1 with unique posttranslational modifications. The epitope is localized to the 246
C
-terminal amino acids within the coiled-coil domain. The 9
C
-terminal residues are known to be critically involved in laminin recognition by integrins.
Prognostic gene signatures for non-small-cell lung cancer
Authors: Boutros Paul C. , Lau Suzanne K. , Pintilie Melania , Liu Ni , Shepherd Frances A. , Der Sandy D. , Tsao Ming-Sound , Penn Linda Z. , Jurisica Igor
Published: February 2009

Resectable non-small-cell lung cancer (NSCLC) patients have poor prognosis, with 30–50% relapsing within 5 years. Current staging criteria do not fully capture the complexity of this disease. Survival could be improved by identification of those early-stage patients who are most likely to benefit from adjuvant therapy. Molecular classification by using mRNA expression profiles has led to multiple, poorly overlapping signatures. We hypothesized that differing statistical methodologies contribute to this lack of overlap. To test this hypothesis, we analyzed our previously published quantitative RT-PCR dataset with a semisupervised method. A 6-gene signature was identified and validated in 4 independent public microarray datasets that represent a range of tumor histologies and stages. This result demonstrated that at least 2 prognostic signatures can be derived from this single dataset. We next estimated the total number of prognostic signatures in this dataset with a 10-million-signature permutation study. Our 6-gene signature was among the top 0.02% of signatures with maximum verifiability, reaffirming its efficacy. Importantly, this analysis identified 1,789 unique signatures, implying that our dataset contains >500,000 verifiable prognostic signatures for NSCLC. This result appears to rationalize the observed lack of overlap among reported NSCLC prognostic signatures.
CHIP regulates leucine-rich repeat kinase-2 ubiquitination, degradation, and toxicity
Authors: Ko Han Seok , Bailey Rachel , Smith Wanli W. , Liu Zhaohui , Shin Joo-Ho , Lee Yun-Il , Zhang Yong-Jie , Jiang Haibing , Ross Christopher A. , Moore Darren J. , Patterson Cam , Petrucelli Leonard , Dawson Ted M. , Dawson Valina L.
Published: February 2009

Mutation in
leucine-rich repeat kinase-2
(
LRRK2
) is the most common cause of late-onset Parkinson's disease (PD). Although most cases of PD are sporadic, some are inherited, including those caused by
LRRK2
mutations. Because these mutations may be associated with a toxic gain of function, controlling the expression of LRRK2 may decrease its cytotoxicity. Here we show that the carboxyl terminus of HSP70-interacting protein (CHIP) binds, ubiquitinates, and promotes the ubiquitin proteasomal degradation of LRRK2. Overexpression of CHIP protects against and knockdown of CHIP exacerbates toxicity mediated by mutant LRRK2. Moreover, HSP90 forms a complex with LRRK2, and inhibition of HSP90 chaperone activity by 17AAG leads to proteasomal degradation of LRRK2, resulting in increased cell viability. Thus, increasing CHIP E3 ligase activity and blocking HSP90 chaperone activity can prevent the deleterious effects of LRRK2. These findings point to potential treatment options for LRRK2-associated PD.
Transforming activity of AML1-ETO is independent of CBFβ and ETO interaction but requires formation of homo-oligomeric complexes
Authors: Kwok Colin , Zeisig Bernd B. , Qiu Jihui , Dong Shuo , So Chi Wai Eric
Published: February 2009

Although both heterodimeric subunits of core binding factors (AML1/RUNX1 and CBFβ) essential for normal hematopoiesis are frequently mutated to form different chimeric fusion proteins in acute leukemia, the underlying molecular mechanisms and structural domains required for cellular transformation remain largely unknown. Despite the critical role of CBFβ for wild-type AML1 function and its direct involvement in chromosomal translocation, we demonstrate that both the expression and interaction with CBFβ are superfluous for AML1-ETO (AE)-mediated transformation of primary hematopoietic cells. Similarly, the hetero-oligomeric interaction with transcriptional repressor ETO family proteins and the highly conserved NHR1 domain in AE fusion are also dispensable for transforming activity. In contrast, AE-mediated transformation is critically dependent on the DNA binding and homo-oligomeric properties of the fusion. Abolishment of homo-oligomerization by a small-molecule inhibitor could specifically suppress AML1 fusion-mediated transformation of primary hematopoietic cells. Together, these results not only identify the essential molecular components but also potential avenues for therapeutic targeting of AE-mediated leukemogenesis.
Phosphorylation of p53 by IκB kinase 2 promotes its degradation by β-TrCP
Authors: Xia Yifeng , Padre Roanna C. , De Mendoza Tatiana Hurtado , Bottero Virginie , Tergaonkar Vinay B. , Verma Inder M.
Published: February 2009

Functional inactivation of p53 and constitutive activation of the NF-κB pathway has been associated with several human cancers. In this study, we show that IκB kinase 2 (IKK2/IKKβ), which is critical for NF-κB activation, also phosphorylates p53. Phosphorylation of p53 at serines 362 and 366 by IKK2 leads to its recruitment to and ubiquitination by β-TrCP1. Degradation of ubiquitinated p53 is independent of Mdm2, because it occurs in both wild-type and Mdm2
−/−
cells. SiRNA-mediated reduction in the levels of β-TrCP1 and other members of the SCF
β−TrCP1
E3 ubiquitin ligase complex or overexpression of a dominant negative form of β-TrCP1 enhances p53 stability. Substitutions at Ser-362 and 366 of p53 by alanines (p53 AA) result in reduced phosphorylation of p53 by IKK2, decreased association with β-TrCP1, and thus increased stability of p53 and expression of p53 target genes such as p21, altering the G1 phase of the cell cycle. Our results identify IKK2 and β-TrCP1 as novel regulators of the p53 pathway and suggest that blocking of IKK2 and β-TrCP1 could be a means of regulating p53 stability and thereby modulating its biological activity.
Integrity of the AID serine-38 phosphorylation site is critical for class switch recombination and somatic hypermutation in mice
Authors: Cheng Hwei-Ling , Vuong Bao Q. , Basu Uttiya , Franklin Andrew , Schwer Bjoern , Astarita Jillian , Phan Ryan T. , Datta Abhishek , Manis John , Alt Frederick W. , Chaudhuri Jayanta
Published: February 2009

Activation-induced cytidine deaminase (AID) is a single-stranded (ss) DNA-specific cytidine deaminase that initiates Ig heavy chain (IgH) class switch recombination (CSR) and Ig somatic hypermutation (SHM) by deaminating cytidines within, respectively, IgH switch (S) regions and Ig variable region (V) exons. AID that is phosphorylated on serine residue 38 interacts with replication protein A (RPA), a ssDNA binding protein, to promote deamination of transcribed double-stranded DNA in vitro, which, along with other evidence, suggests that AID may similarly gain access to transcribed S regions and V exons in vivo. However, the physiological role of AID phosphorylation at serine residue 38 (S38), and even the requirement for the S38 residue, with respect to CSR or SHM has been debated. To address this issue, we used gene targeting to generate an endogenous mouse
AID
locus that produces AID in which S38 is substituted with alanine (AID
S38A
), a mutant form of AID that retains similar catalytic activity on ssDNA as WT AID (AID
WT
). B cells homozygous for the AID
S38A
mutation show substantially impaired CSR and SHM, correlating with inability of AID
S38A
to interact with endogenous RPA. Moreover, mice haploinsufficient for AID
S38A
have even more severely impaired CSR when compared with mice haploinsufficient for AID
WT
, with CSR levels reduced to nearly background levels. These results unequivocally demonstrate that integrity of the AID S38 phosphorylation site is required for normal CSR and SHM in mice and strongly support a role for AID phosphorylation at S38 and RPA interaction in regulating CSR and SHM.
Urea, but not guanidinium, destabilizes proteins by forming hydrogen bonds to the peptide group
Authors: Lim Woon Ki , Rösgen Jörg , Englander S. Walter
Published: February 2009

The mechanism by which urea and guanidinium destabilize protein structure is controversial. We tested the possibility that these denaturants form hydrogen bonds with peptide groups by measuring their ability to block acid- and base-catalyzed peptide hydrogen exchange. The peptide hydrogen bonding found appears sufficient to explain the thermodynamic denaturing effect of urea. Results for guanidinium, however, are contrary to the expectation that it might H-bond. Evidently, urea and guanidinium, although structurally similar, denature proteins by different mechanisms.
A role for epithelial-mesenchymal transition in the etiology of benign prostatic hyperplasia
Authors: Alonso-Magdalena Paloma , Brössner Clemens , Reiner Angelika , Cheng Guojun , Sugiyama Nobuhiro , Warner Margaret , Gustafsson Jan-Åke
Published: February 2009

Benign prostatic hyperplasia (BPH) is usually described as a pathological proliferation of prostatic fibroblasts/myofibroblasts and epithelial cells. In the present study of BPH samples, we have made a morphological and immunohistochemical study of BPH prostatic sections using markers of proliferation, apoptosis, hormone receptors, and TGF-β signaling. We found no evidence of proliferation in the stroma but in the epithelium of some ducts; 0.7% of the basal and 0.4% of luminal cells were positive for Ki67 and PCNA. Androgen receptor and estrogen receptor beta (ERβ)1 and ERβcx were abundant in both stromal and epithelial compartments but cells expressing ERα were very rare. What was very common in all BPH samples was the following: (
i
) regions of the ductal epithelium where the epithelial cells did not express E-cadherin, had lost their polarization, and become spindle shaped (the nuclei of these cells were strongly positive for pSmad 3 and Snail); and (
ii
) regions where the walls of the blood vessels were extremely thick and there was loss of endothelial layer. Loss of E-cadherin, increased pSmad 3, and high expression of Snail are all characteristic of epithelial-mesenchymal transition (EMT). We conclude that BPH is not a disease of prostatic stromal proliferation but rather of accumulation of mesenchymal-like cells derived from the prostatic epithelium and the endothelium. TGF-β is thought to play a key role in EMT. Our data suggests that TGF-β/Smad should be considered as targets for treatment of BPH.
Dynamic regulation of mitochondrial function by glucocorticoids
Authors: Du Jing , Wang Yun , Hunter Richard , Wei Yanling , Blumenthal Rayah , Falke Cynthia , Khairova Rushaniya , Zhou Rulun , Yuan Peixiong , Machado-Vieira Rodrigo , McEwen Bruce S. , Manji Husseini K.
Published: February 2009

Glucocorticoids play an important biphasic role in modulating neural plasticity; low doses enhance neural plasticity and spatial memory behavior, whereas chronic, higher doses produce inhibition. We found that 3 independent measures of mitochondrial function—mitochondrial oxidation, membrane potential, and mitochondrial calcium holding capacity—were regulated by long-term corticosterone (CORT) treatment in an inverted “U”-shape. This regulation of mitochondrial function by CORT correlated with neuroprotection; that is, treatment with low doses of CORT had a neuroprotective effect, whereas treatment with high doses of CORT enhanced kainic acid (KA)-induced toxicity of cortical neurons. We then undertook experiments to elucidate the mechanisms underlying these biphasic effects and found that glucocorticoid receptors (GRs) formed a complex with the anti-apoptotic protein Bcl-2 in response to CORT treatment and translocated with Bcl-2 into mitochondria after acute treatment with low or high doses of CORT in primary cortical neurons. However, after 3 days of treatment, high, but not low, doses of CORT resulted in decreased GR and Bcl-2 levels in mitochondria. As with the in vitro studies, Bcl-2 levels in the mitochondria of the prefrontal cortex were significantly decreased, along with GR levels, after long-term treatment with high-dose CORT in vivo. These findings have the potential to contribute to a more complete understanding of the mechanisms by which glucocorticoids and chronic stress regulate cellular plasticity and resilience and to inform the future development of improved therapeutics.
Carboxy-terminal domain of AID required for its mRNA complex formation in vivo
Authors: Nonaka Taichiro , Doi Tomomitsu , Toyoshima Takae , Muramatsu Masamichi , Honjo Tasuku , Kinoshita Kazuo
Published: February 2009

Activation-induced cytidine deaminase (AID) is essential for the class switch recombination (CSR) and somatic hypermutation (SHM) of Ig genes. Originally, AID was postulated to be an RNA-editing enzyme, because of its structural homology with a known RNA-editing enzyme, APOBEC1. In support of this idea, AID shares many of the properties of RNA-editing enzymes, including nucleocytoplasmic shuttling and a dependency on de novo protein synthesis. However, it has not been shown whether AID recognizes a specific mRNA and edits it to generate an enzyme involved in CSR or SHM. Here, we examined the association between AID and polyadenylated [poly(A)
+
] RNA in vivo, using UV cross-linking coupled with a poly(A) capture method that relies on biotinylated oligo(dT) and streptavidin-conjugated beads. We found that both exogenous AID expressed in transfected CH12 cells and endogenous AID expressed in BL2 cells were associated with poly(A)
+
RNA. Similar protein-poly(A)
+
RNA complexes were formed by APOBEC1 and APOBEC3G. However, the interactions of all of these cytidine deaminase family members, including AID, with poly(A)
+
RNA were indirect. This was expected for APOBEC1, which is known to act through an RNA-interacting cofactor, APOBEC1 complementation factor (ACF). In addition, the carboxy-terminal region of AID, which is essential for class switching, was also required for its interaction with poly(A)
+
RNA. These results suggest that the CSR activity of AID requires an ACF-like cofactor that specifically interacts with the carboxy-terminal domain of AID.
Anti-CTLA-4 therapy results in higher CD4
+
ICOS
hi
T cell frequency and IFN-γ levels in both nonmalignant and malignant prostate tissues
Authors: Chen Hong , Liakou Chrysoula I. , Kamat Ashish , Pettaway Curtis , Ward John F. , Tang Derek Ng , Sun Jingjing , Jungbluth Achim A. , Troncoso Patricia , Logothetis Christopher , Sharma Padmanee
Published: February 2009

Cytotoxic lymphocyte antigen-4 (CTLA-4) blockade is an active immunotherapeutic strategy that is currently in clinical trials in cancer. There are several ongoing trials of anti-CTLA-4 in the metastatic setting of prostate cancer patients with reported clinical responses consisting of decreases in the prostate specific antigen (PSA) tumor marker for some patients. Immunologic markers that correlate with these clinical responses are necessary to guide further development of anti-CTLA-4 therapy in the treatment of cancer patients. We recently reported that CD4
+
inducible co-stimulator (ICOS)
hi
T cells that produce interferon-γ (IFN-γ) are increased in the peripheral blood and tumor tissues of bladder cancer patients treated with anti-CTLA-4 antibody. Here we present data from the same clinical trial in bladder cancer patients demonstrating a higher frequency of CD4
+
ICOS
hi
T cells and IFN-γ mRNA levels in nonmalignant prostate tissues and incidental prostate tumor tissues removed at the time of radical cystoprostatectomy. Our data suggest immunologic markers that can be used to monitor prostate cancer patients who receive anti-CTLA-4 therapy and indicate that the immunologic impact of anti-CTLA-4 antibody can occur in both tumor and nonmalignant tissues. These data should be taken into consideration for evaluation of efficacy as well as immune-related adverse events associated with anti-CTLA-4 therapy.
Mice with mutations of
Dock7
have generalized hypopigmentation and white-spotting but show normal neurological function
Authors: Blasius Amanda L. , Brandl Katharina , Crozat Karine , Xia Yu , Khovananth Kevin , Krebs Philippe , Smart Nora G. , Zampolli Antonella , Ruggeri Zaverio M. , Beutler Bruce A.
Published: February 2009

The classical recessive coat color mutation
misty
(
m
) arose spontaneously on the DBA/J background and causes generalized hypopigmentation and localized white-spotting in mice, with a lack of pigment on the belly, tail tip, and paws. Here we describe
moonlight
(
mnlt
), a second hypopigmentation and white-spotting mutation identified on the C57BL/6J background, which yields a phenotypic copy of
m
/
m
coat color traits. We demonstrate that the 2 mutations are allelic.
m
/
m
and
mnlt
/
mnlt
phenotypes both result from mutations that truncate the dedicator of cytokinesis 7 protein (DOCK7), a widely expressed Rho family guanine nucleotide exchange factor. Although
Dock7
is transcribed at high levels in the developing brain and has been implicated in both axon development and myelination by in vitro studies, we find no requirement for DOCK7 in neurobehavioral function in vivo. However, DOCK7 has non-redundant role(s) related to the distribution and function of dermal and follicular melanocytes.
Loss of p53 enhances catalytic activity of IKKβ through O-linked β-N-acetyl glucosamine modification
Authors: Kawauchi Keiko , Araki Keigo , Tobiume Kei , Tanaka Nobuyuki
Published: February 2009

The IκB kinase (IKK)-NF-κB pathway plays a critical role in oncogenesis. Recently, we have shown that p53 regulates glucose metabolism through the IKK-NF-κB pathway and that, in the absence of p53, the positive feedback loop between IKK-NF-κB and glycolysis has an integral role in oncogene-induced cell transformation. Here, we demonstrate that IKKβ, a component of the IKK complex, was constitutively modified with O-linked β-N-acetyl glucosamine (O-GlcNAc) in both
p53
-deficient mouse embryonic fibroblasts (MEFs) and transformed human fibroblasts. In
p53
-deficient cells, the O-GlcNAcylated IKKβ and the activating phosphorylation of IKK were decreased by p65/NF-κB knockdown or glucose depletion. We also found that high glucose induced the O-GlcNAcylation of IKKβ and sustained the TNFα-dependent IKKβ activity. Moreover, the O-GlcNAcase inhibitor streptozotocin intensified O-GlcNAcylation and concomitant activating phosphorylation of IKKβ. Mutational analysis revealed that O-GlcNAcylation of IKKβ occurred at Ser 733 in the C-terminal domain, which was identified as an inactivating phosphorylation site, suggesting that IKKβ O-GlcNAcylation regulates its catalytic activity. Taken together, we propose a novel mechanism for the enhancement of NF-κB activity by loss of p53, which evokes positive feedback regulation from enhanced glucose metabolism to IKK in oncogenesis.
Further evidence for involvement of a noncanonical function of uracil DNA glycosylase in class switch recombination
Authors: Begum Nasim A. , Stanlie Andre , Doi Tomomitsu , Sasaki Yoko , Jin Hai Wei , Kim Yong Sung , Nagaoka Hitoshi , Honjo Tasuku
Published: February 2009

Activation-induced cytidine deaminase (AID) introduces DNA cleavage in the Ig gene locus to initiate somatic hypermutation (SHM) and class switch recombination (CSR) in B cells. The DNA deamination model assumes that AID deaminates cytidine (C) on DNA and generates uridine (U), resulting in DNA cleavage after removal of U by uracil DNA glycosylase (UNG). Although UNG deficiency reduces CSR efficiency to one tenth, we reported that catalytically inactive mutants of UNG were fully proficient in CSR and that several mutants at noncatalytic sites lost CSR activity, indicating that enzymatic activity of UNG is not required for CSR. In this report we show that CSR activity by many UNG mutants critically depends on its N-terminal domain, irrespective of their enzymatic activities. Dissociation of the catalytic and CSR activity was also found in another UNG family member, SMUG1, and its mutants. We also show that Ugi, a specific peptide inhibitor of UNG, inhibits CSR without reducing DNA cleavage of the S (switch) region, confirming dispensability of UNG in DNA cleavage in CSR. It is therefore likely that UNG is involved in a repair step after DNA cleavage in CSR. Furthermore, requirement of the N terminus but not enzymatic activity of UNG mutants for CSR indicates that the UNG protein structure is critical. The present findings support our earlier proposal that CSR depends on a noncanonical function of the UNG protein (e.g., as a scaffold for repair enzymes) that might be required for the recombination reaction after DNA cleavage.
Nrf2-mediated neuroprotection in the MPTP mouse model of Parkinson's disease: Critical role for the astrocyte
Authors: Chen Pei-Chun , Vargas Marcelo R. , Pani Amar K. , Smeyne Richard J. , Johnson Delinda A. , Kan Yuet Wai , Johnson Jeffrey A.
Published: February 2009

Oxidative stress has been implicated in the etiology of Parkinson's disease (PD) and in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) animal model of PD. It is known that under conditions of oxidative stress, the transcription factor NF-E2-related factor (Nrf2) binds to antioxidant response element (ARE) to induce antioxidant and phase II detoxification enzymes. To investigate the role of Nrf2 in the process of MPTP-induced toxicity, mice expressing the human placental alkaline phosphatase (hPAP) gene driven by a promoter containing a core ARE sequence (ARE-hPAP) were used. ARE-hPAP mice were injected (30 mg/kg) once per day for 5 days and killed 7 days after the last MPTP injection. In response to this design, ARE-dependent gene expression was decreased in striatum whereas it was increased in substantia nigra. The same MPTP protocol was applied in Nrf2+/+and Nrf2−/−mice; Nrf2 deficiency increases MPTP sensitivity. Furthermore, we evaluated the potential for astrocytic Nrf2 overexpression to protect from MPTP toxicity. Transgenic mice with Nrf2 under control of the astrocyte-specific promoter for the glial fribillary acidic protein (GFAP-Nrf2) on both a Nrf2+/+and Nrf2−/−background were administered MPTP. In the latter case, only the astrocytes expressed Nrf2. Independent of background, MPTP-mediated toxicity was abolished in GFAP-Nrf2 mice. These striking results indicate that Nrf2 expression restricted to astrocytes is sufficient to protect against MPTP and astrocytic modulation of the Nrf2-ARE pathway is a promising target for therapeutics aimed at reducing or preventing neuronal death in PD.
Antioxidant and Preventive Effects of Extract from Nymphaea candida Flower on In Vitro Immunological Liver Injury of Rat Primary Hepatocyte Cultures
Authors: Zhao Jun , Liu Tao , Ma Long , Yan Ming , Gu Zhengyi , Huang Yi , Xu Fang , Zhao Yu
Published: February 2009

Nymphaea candida is traditional Uighur medicine that is commonly used to treat head pains, cough, hepatitis and hypertension in Xinjiang of China. In this article, the extract of N. candida was measured for antioxidant activity, using 1,1‐diphenyl‐2‐picrylhydrazyl (DPPH) radicals scavenging assay and reducing power determination, and compared with those of the positive controls of butylated hydroxytoluene (BHT) and gallic acid (GA). The active extract was further purified by liquid‐liquid partition to afford four fractions, of which the ethyl acetate‐soluble (EA) fraction (NCE) exhibited the strongest antioxidant capacity with IC50 value of 12.6 μg/mL for DPPH. Thirteen phenolic compounds were isolated from this fraction, and they all showed significant antioxidant activities in DPPH model system. Furthermore, NCE showed potent antioxidant capacity with IC50 value of 59.32 μg/mL, 24.48 μg/mL and 86.85 μg/mL, for , ·OH and H2O2 radicals, respectively. Moreover, NCE on BCG plus LPS‐induced immunological liver injury was evaluated using primary cultured rat hepatocytes. NCE produced significant hepatoprotective effects as evidenced by decreased supernatant enzyme activities (AST—aspartate transaminase, P <  .01; ALT—alanine transferase, P <  .01) and nitric oxide (NO, P <  .01) production. These results revealed the in vitro antioxidant and hepatoprotective activities of NCE against immunological liver injury. Further investigations are necessary to verify these activities in vivo.
Distinct Amino Acids of the Phytophthora infestans Effector AVR3a Condition Activation of R3a Hypersensitivity and Suppression of Cell Death
Authors: Bos Jorunn I. B. , Chaparro-Garcia Angela , Quesada-Ocampo Lina M. , Gardener Brian B. McSpadden , Kamoun Sophien
Published: February 2009

The AVR3a protein of Phytophthora infestans is a polymorphic member of the RXLR class of cytoplasmic effectors with dual functions. AVR3aKI but not AVR3aEM activates innate immunity triggered by the potato resistance protein R3a and is a strong suppressor of the cell-death response induced by INF1 elicitin, a secreted P. infestans protein that has features of pathogen-associated molecular patterns. To gain insights into the molecular basis of AVR3a activities, we performed structure-function analyses of both AVR3a forms. We utilized saturated high-throughput mutant screens to identify amino acids important for R3a activation. Of 6,500 AVR3aEM clones tested, we identified 136 AVR3aEM mutant clones that gained the ability to induce R3a hypersensitivity. Fifteen amino-acid sites were affected in this set of mutant clones. Most of these mutants did not suppress cell death at a level similar to that of AVR3aKI. A similar loss-of-function screen of 4,500 AVR3aKI clones identified only 13 mutants with altered activity. These results point to models in which AVR3a functions by interacting with one or more host proteins and are not consistent with the recognition of AVR3a through an enzymatic activity. The identification of mutants that gain R3a activation but not cell-death suppression activity suggests that distinct amino acids condition the two AVR3a effector activities.
First Report of Smut on Imperata cylindrica Caused by Sporisorium schweinfurthianum in South Africa
Authors: Wood A. R. , den Breeÿen A. , Beed F.
Published: February 2009

Imperata cylindrica (L.) Raeusch. (Poaceae) is indigenous to the old world but is a problem weed in tropical areas throughout the world (1). A smut fungus was observed frequently on this grass at a single site near Pretoria (25°44′19″S, 28°15′39″E), South Africa during April of 2006. On the basis of the following characteristics, it was identified as Sporisorium schweinfurthianum (Thüm.) K. Vánky (2). Panicles were systemically infected and all ovaries in infected inflorescences were replaced by spores. Spores were globose or subglobose, brown, 10 to 14 × 9 to 12 μm (average 11.2 × 9.8 μm; n = 25), wall 1 μm thick, and finely verruculose. Hyaline, thin-walled sterile cells were present. This identification was confirmed by K. Vánky (personal communication to A. R. Wood). To our knowledge, this is the first report of this smut species from southern Africa. A voucher specimen has been deposited in the South African National Collection of Fungi, ARC-Plant Protection Research Institute (PREM 59895). To test pathogenicity, soil in 15 pots with individual 1-month-old seedlings was drenched with an aqueous suspension of 1 × 108 spores ml–1 amended with 0.1% Tween 80. Before treatment, the pots were placed on pot trays and remained immersed in the spore suspension in the trays at 28°C (relative humidity <80%) for 24 h. To maintain the spore concentration in the soil, the pots were not watered until 7 days after inoculation. Distilled water amended with 0.1% Tween 80 was applied as control treatments to a further 15 pots with plants. Five of the treated plants produced panicles within 4 months of inoculation. Of these, all the ovaries of four emerging inflorescences were completely replaced with a brown, powdery mass of teliospores. No smutted panicles developed on the control plants. This smut fungus may have potential as a classical biological control agent for use against I. cylindrica by reducing dispersal by seed.

References: (1) L. G. Holm et al. The World's Worst Weeds: Distribution and Biology. University Press of Hawaii. Honolulu, 1977. (2) K. Vánky. Australas. Plant Pathol. 29:155, 2000.
Development and Characterization of Expressed Sequence Tag-Derived Microsatellite Markers for the Wheat Stem Rust Fungus Puccinia graminis f. sp. tritici
Authors: Zhong S. , Leng Y. , Friesen T. L. , Faris J. D. , Szabo L. J.
Published: February 2009

Puccinia graminis f. sp. tritici is the causal agent of stem rust disease in wheat. The rust fungus has caused devastating disease epidemics throughout history and is still posing a potential threat to wheat production in some regions of the world due to the appearance of new races. To develop microsatellite or simple sequence repeat (SSR) markers for use in population genetics studies, a total of 60,579 expressed sequence tag (EST) sequences (reads) generated from P. graminis f. sp. tritici were screened for tandemly repeated di- and tri-nucleotide units using a bioinformatics approach and 708 unisequences containing putative SSR loci with six or more repeat units were identified. Flanking primers were designed for 384 unique SSR loci, which mapped to different locations of the draft genome sequence of the fungus. Of the 384 primer pairs tested, 72 EST-SSR markers were eventually developed, which showed polymorphism among 19 isolates of P. graminis f. sp. tritici and 4 isolates of P. graminis f. sp. secalis evaluated. Thirty-two of the SSR loci were also evaluated in three other rust fungi (P. triticina, P. hordei, and P. coronata f. sp. hordei) for cross-species transferability. These SSR markers derived from ESTs will be useful for characterization of population structures and for gene mapping in P. graminis.
Secreted indole serves as a signal for expression of type III secretion system translocators in enterohaemorrhagic Escherichia coli O157 : H7
Authors: Hirakawa Hidetada , Kodama Toshio , Takumi-Kobayashi Asuka , Honda Takeshi , Yamaguchi Akihito
Published: February 2009

Indole is produced by tryptophanase during growth of enteric bacteria and accumulates in the culture medium. The physiological role of indole production is poorly understood. We discovered that enterohaemorrhagic Escherichia coli (EHEC) O157 : H7 with a tnaA deletion has decreased secretion of EspA and EspB via the type III secretion system and as a result there is reduced formation of attaching and effacing (A/E) lesions in HeLa cells. Addition of indole restored and enhanced secretion of EspA and EspB and formation of A/E lesions by the tnaA deletion mutant EHEC. Indole addition moderately increased the promoter activity of LEE4 genes, including espA and espB, in the locus of enterocyte effacement. Thus in EHEC indole can serve to signal EspA and EspB expression and secretion and stimulate the ability of EHEC to form A/E lesions on human cells.
Identification of genes required for different stages of dendritic swarming in Bacillus subtilis, with a novel role for phrC
Authors: Hamze Kassem , Julkowska Daria , Autret Sabine , Hinc Krzysztof , Nagorska Krzysztofa , Sekowska Agnieszka , Holland I. Barry , Séror Simone J.
Published: February 2009

Highly branched dendritic swarming ofB. subtilison synthetic B-medium involves a developmental-like process that is absolutely dependent on flagella and surfactin secretion. In order to identify new swarming genes, we targeted the two-component ComPA signalling pathway and associated global regulators. In liquid cultures, the histidine kinase ComP, and the response regulator ComA, respond to secreted pheromones ComX and CSF (encoded byphrC) in order to control production of surfactin synthases and ComS (competence regulator). In this study, for what is believed to be the first time, we established that distinct early stages of dendritic swarming can be clearly defined, and that they are amenable to genetic analysis. In a mutational analysis producing several mutants with distinctive phenotypes, we were able to assign the genessfp(activation of surfactin synthases),comA, abrBandcodY(global regulators),hag(flagellin),mecAandyvzB(hag-like),and swrB(motility), to the different swarming stages. Surprisingly, mutations in genescomPX, comQ, comS,rapCandoppD, which are normally indispensable for import of CSF, had only modest effects, if any, on swarming and surfactin production. Therefore, during dendritic swarming, surfactin synthesis is apparently subject to novel regulation that is largely independent of the ComXP pathway; we discuss possible alternative mechanisms for drivingsrfABCDtranscription. We showed that thephrCmutant, largely independent of any effect on surfactin production, was also, nevertheless, blocked early in swarming, forming stunted dendrites, with abnormal dendrite initiation morphology. In a mixed swarm co-inoculated withphrC sfp+andphrC+sfp(GFP), an apparently normal swarm was produced. In fact, while initiation of all dendrites was of the abnormalphrCtype, these were predominantly populated bysfpcells, which migrated faster than thephrCcells. This and other results indicated a specific migration defect in thephrCmutant that could not betrans-complemented by CSF in a mixed swarm. CSF is the C-terminal pentapeptide of the surface-exposed PhrC pre-peptide and we propose that the residual PhrC 35 aa residue peptide anchored in the exterior of the cytoplasmic membrane has an apparently novel extracellular role in swarming.
Probiotics prevent enterohaemorrhagic Escherichia coli O157 : H7-mediated inhibition of interferon-γ-induced tyrosine phosphorylation of STAT-1
Authors: Jandu Narveen , Zeng Zoë Jingjing , Johnson-Henry Kathene C. , Sherman Philip M.
Published: February 2009

Enterohaemorrhagic Escherichia coli (EHEC) O157 : H7 inhibits interferon (IFN)-γ-stimulated tyrosine phosphorylation of signal transducer and activator of transcription (STAT)-1 in epithelial cells. We determined the effects of probiotics on EHEC-mediated disruption of IFN-γ-stimulated STAT-1 activation in epithelial cell lines. Confluent Intestine 407, HEp-2 and Caco-2 epithelial cells were pre-treated (3 h) with either probiotics or surface-layer proteins derived from Lactobacillus helveticus R0052 prior to infection with EHEC O157 : H7 strain CL56 (m.o.i. 100 : 1, 6 h, 37 °C in 5 % CO2). Subsequently, cells were washed and stimulated with human recombinant IFN-γ (50 ng ml−1, 0.5 h, 37 °C) followed by whole-cell protein extraction and immunoblotting for tyrosine-phosphorylated STAT-1. Relative to uninfected cells, STAT-1-activation was reduced after EHEC O157 : H7 infection. Pre-incubation with the probiotic L. helveticus R0052 followed by EHEC infection abrogated pathogen-mediated disruption of IFN-γ–STAT-1 signalling. As determined using Transwell inserts, probiotic-mediated protection was independent of epithelial cell contact. In contrast, pre-incubation with boiled L. helveticus R0052, an equal concentration of viable Lactobacillus rhamnosus R0011, or surface-layer proteins (0.14 mg ml−1) did not restore STAT-1 signalling in EHEC-infected cells. The viable probiotic agent L. helveticus R0052 prevented EHEC O157 : H7-mediated subversion of epithelial cell signal transduction responses.
Differences between human subjects in the composition of the faecal bacterial community and faecal metabolism of linoleic acid
Authors: Devillard Estelle , McIntosh Freda M. , Paillard Delphine , Thomas Nadine A. , Shingfield Kevin J. , Wallace R. John
Published: February 2009

Conjugated linoleic acid (CLA) is formed from linoleic acid (LA;cis-9,cis-12-18 : 2) by intestinal bacteria. Different CLA isomers have different implications for human health. The aim of this study was to investigate LA metabolism and the CLA isomers formed in two individuals (V1 and V2) with different faecal metabolic characteristics, and to compare fatty acid metabolism with the microbial community composition. LA incubated with faecal samples was metabolized at similar rates with both subjects, but the products were different. LA was metabolized extensively to stearic acid (SA; 18 : 0) in V1, with minor accumulation of CLA and more rapid accumulation of vaccenic acid (VA;trans-11-18 : 1). CLA accumulation at 4 h was almost tenfold higher with V2, and little SA was formed. At least 12 different isomers of CLA were produced from LA by the colonic bacteria from the two individuals. The predominant (>75 %) CLA isomer in V1 was rumenic acid (RA;cis-9,trans-11-18 : 2), whereas the concentrations of RA andtrans-10,cis-12-18 : 2 were similar with V2. Propionate and butyrate proportions in short-chain fatty acids were higher in V1. A 16S rRNA clone library from V1 contained mainlyBacteroidetes(54 % of clones), whereasFirmicutes(66 % of clones) predominated in V2. Both samples were devoid of bacteria related toClostridium proteoclasticum, the only gut bacterium known to metabolize VA to SA. Thus, the CLA formed in the intestine of different individuals may differ according to their resident microbiota, with possibly important implications with respect to gut health.
Plasmid diversity in Vibrio vulnificus biotypes
Authors: Roig Francisco J. , Amaro Carmen
Published: February 2009

Vibrio vulnificus is a heterogeneous bacterial species that can be virulent for humans and fish. Virulence in fish seems to rely on a recently described plasmid that can be transmitted between strains, aided by a conjugative plasmid. The main objective of this work was to analyse the plasmid content of a wide collection of strains from the three biotypes of the species, as well as to identify putative conjugative and virulence plasmids by means of Southern hybridization with specific probes and sequence analysis of selected gene markers. We found 28 different plasmid profiles in a total of 112 strains, which were relatively biotype- or serovar-specific. Biotype 1 lacked high-molecular-mass plasmids, with the exception of a putative conjugative plasmid of 48 kb that was present in 42.8 % of clinical and environmental strains isolated worldwide. All biotype 2 strains possessed the virulence plasmid, whose molecular mass ranged between 68 and 70 kb, and 89.65 % of these strains also had a putative conjugative plasmid with a molecular size of 52–56 kb. Finally, a 48 kb putative conjugative plasmid was present in all biotype 3 strains. Data from partial sequencing of traD, traI and the whole vep07 (a recently described plasmid-borne virulence gene) from a selection of strains suggest that the plasmids of 48–56 kb probably belong to the same family of F-plasmids as pYJ016 and that the gene vep07 is absolutely essential for fish virulence. Additional cryptic plasmids of low molecular mass were present in the three biotypes. In conclusion, plasmids are widespread among V. vulnificus species and could contribute substantially to genetic plasticity of the species.