bluuebunny/crossref_metadata_2025 huggingface dataset:
Structural evolution of p53, p63, and p73: Implication for heterotetramer formation
Authors: Joerger Andreas C. , Rajagopalan Sridharan , Natan Eviatar , Veprintsev Dmitry B. , Robinson Carol V. , Fersht Alan R.
Published: October 2009

Oligomerization of members of the p53 family of transcription factors (p53, p63, and p73) is essential for their distinct functions in cell-cycle control and development. To elucidate the molecular basis for tetramer formation of the various family members, we solved the crystal structure of the human p73 tetramerization domain (residues 351–399). Similarly to the canonical p53 tetramer, p73 forms a tetramer with D
2
symmetry that can be described as a dimer of dimers. The most striking difference between the p53 and p73 tetramerization domain is the presence of an additional C-terminal helix in p73. This helix, which is conserved in p63, is essential for stabilizing the overall architecture of the tetramer, as evidenced by the different oligomeric structures observed for a shortened variant lacking this helix. The helices act as clamps, wrapping around the neighboring dimer and holding it in place. In addition, we show by mass spectrometry that the tetramerization domains of p63 and p73, but not p53, fully exchange, with different mixed tetramers present at equilibrium, albeit at a relatively slow rate. Taken together, these data provide intriguing insights into the divergent evolution of the oligomerization domain within the p53 family, from the ancestral p63/p73-like protein toward smaller, less promiscuous monomeric building blocks in human p53, allowing functional separation of the p53 pathway from that of its family members.
Transgenic songbirds offer an opportunity to develop a genetic model for vocal learning
Authors: Agate R. J. , Scott B. B. , Haripal B. , Lois C. , Nottebohm F.
Published: October 2009

Zebra finches are widely used for studying the basic biology of vocal learning. The inability to introduce genetic modifications in these animals has substantially limited studies on the molecular biology of this behavior, however. We used an HIV-based lentivirus to produce germline transgenic zebra finches. The lentivirus encoded the GFP regulated by the human ubiquitin-C promoter [Lois C, Hong EJ, Pease S, Brown EJ, Baltimore D (2002)
Science
295:868–872], which is active in a wide variety of cells. The virus was injected into the very early embryo (blastodisc stage) to target the primordial germline cells that later give rise to sperm and eggs. A total of 265 fertile eggs were injected with virus, and 35 hatched (13%); 23 of these potential founders (F0) were bred, and three (13%) produced germline transgenic hatchlings that expressed the GFP protein (F1). Two of these three founders (F0) have produced transgenic young at a rate of 12% and the third at a rate of 6%. Furthermore, two of the F1 generation transgenics have since reproduced, one having five offspring (all GFP positive) and the other four offsping (one GFP positive).
The Exomer Coat Complex Transports Fus1p to the Plasma Membrane via a Novel Plasma Membrane Sorting Signal in Yeast
Authors: Barfield Robyn M. , Fromme J. Christopher , Schekman Randy
Published: October 2009

Sorting of transmembrane cargo proteins to different cellular compartments is mediated by sorting signals that are recognized by coat proteins involved in vesicle biogenesis. We have identified a sorting signal in the yeast cell fusion protein Fus1p that is required for its transport from the trans-Golgi compartment to the plasma membrane. Transport of Fus1p from the trans-Golgi to the cell surface is dependent on Chs5p, a component of the multisubunit exomer complex. We show that Fus1p transport is also dependent on the exomer components Bch1p and Bud7p. Disruption of the clathrin adaptor protein complex 1 (AP-1) restores Fus1p localization to the cell surface in the absence of exomer, possibly by promoting an alternate, exomer-independent route of transport. Mutation of an IXTPK sequence in the cytosolic tail of Fus1p abolishes its physical interaction with Chs5p, results in mislocalization of Fus1p, and therefore causes a cell fusion defect. These defects are suppressed by disruption of AP-1. We suggest that IXTPK comprises a novel sorting signal that is recognized and bound by exomer leading to the capture of Fus1p into coated vesicles en route to the cell surface.
Brd4 Marks Select Genes on Mitotic Chromatin and Directs Postmitotic Transcription
Authors: Dey Anup , Nishiyama Akira , Karpova Tatiana , McNally James , Ozato Keiko
Published: October 2009

On entry into mitosis, many transcription factors dissociate from chromatin, resulting in global transcriptional shutdown. During mitosis, some genes are marked to ensure the inheritance of their expression in the next generation of cells. The nature of mitotic gene marking, however, has been obscure. Brd4 is a double bromodomain protein that localizes to chromosomes during mitosis and is implicated in holding mitotic memory. In interphase, Brd4 interacts with P-TEFb and functions as a global transcriptional coactivator. We found that throughout mitosis, Brd4 remained bound to the transcription start sites of many M/G1 genes that are programmed to be expressed at the end of, or immediately after mitosis. In contrast, Brd4 did not bind to genes that are expressed at later phases of cell cycle. Brd4 binding to M/G1 genes increased at telophase, the end phase of mitosis, coinciding with increased acetylation of histone H3 and H4 in these genes. Increased Brd4 binding was accompanied by the recruitment of P-TEFb and de novo M/G1 gene transcription, the events impaired in Brd4 knockdown cells. In sum, Brd4 marks M/G1 genes for transcriptional memory during mitosis, and upon exiting mitosis, this mark acts as a signal for initiating their prompt transcription in daughter cells.
Neogenin Regulates Skeletal Myofiber Size and Focal Adhesion Kinase and Extracellular Signal-regulated Kinase Activities In Vivo and In Vitro
Authors: Bae Gyu-Un , Yang Youn-Joo , Jiang Guoying , Hong Mingi , Lee Hye-Jin , Tessier-Lavigne Marc , Kang Jong-Sun , Krauss Robert S.
Published: October 2009

A variety of signaling pathways participate in the development of skeletal muscle, but the extracellular cues that regulate such pathways in myofiber formation are not well understood. Neogenin is a receptor for ligands of the netrin and repulsive guidance molecule (RGM) families involved in axon guidance. We reported previously that neogenin promoted myotube formation by C2C12 myoblasts in vitro and that the related protein Cdo (also Cdon) was a potential neogenin coreceptor in myoblasts. We report here that mice homozygous for a gene-trap mutation in the Neo1 locus (encoding neogenin) develop myotomes normally but have small myofibers at embryonic day 18.5 and at 3 wk of age. Similarly, cultured myoblasts derived from such animals form smaller myotubes with fewer nuclei than myoblasts from control animals. These in vivo and in vitro defects are associated with low levels of the activated forms of focal adhesion kinase (FAK) and extracellular signal-regulated kinase (ERK), both known to be involved in myotube formation, and inefficient expression of certain muscle-specific proteins. Recombinant netrin-2 activates FAK and ERK in cultured myoblasts in a neogenin- and Cdo-dependent manner, whereas recombinant RGMc displays lesser ability to activate these kinases. Together, netrin-neogenin signaling is an important extracellular cue in regulation of myogenic differentiation and myofiber size.
Polyamines Regulate c-Myc Translation through Chk2-dependent HuR Phosphorylation
Authors: Liu Lan , Rao Jaladanki N. , Zou Tongtong , Xiao Lan , Wang Peng-Yuan , Turner Douglas J. , Gorospe Myriam , Wang Jian-Ying
Published: October 2009

All mammalian cells depend on polyamines for normal growth and proliferation, but the exact roles of polyamines at the molecular level remain largely unknown. The RNA-binding protein HuR modulates the stability and translation of many target mRNAs. Here, we show that in rat intestinal epithelial cells (IECs), polyamines enhanced HuR association with the 3′-untranslated region of the c-Myc mRNA by increasing HuR phosphorylation by Chk2, in turn promoting c-Myc translation. Depletion of cellular polyamines inhibited Chk2 and reduced the affinity of HuR for c-Myc mRNA; these effects were completely reversed by addition of the polyamine putrescine or by Chk2 overexpression. In cells with high content of cellular polyamines, HuR silencing or Chk2 silencing reduced c-Myc translation and c-Myc expression levels. Our findings demonstrate that polyamines regulate c-Myc translation in IECs through HuR phosphorylation by Chk2 and provide new insight into the molecular functions of cellular polyamines.
A Mutation Associated with CMT2A Neuropathy Causes Defects in Fzo1 GTP Hydrolysis, Ubiquitylation, and Protein Turnover
Authors: Amiott Elizabeth A. , Cohen Mickael M. , Saint-Georges Yann , Weissman Allan M. , Shaw Janet M.
Published: October 2009

Charcot-Marie-Tooth disease type 2A (CMT2A) is caused by mutations in the gene MFN2 and is one of the most common inherited peripheral neuropathies. Mfn2 is one of two mammalian mitofusin GTPases that promote mitochondrial fusion and maintain organelle integrity. It is not known how mitofusin mutations cause axonal degeneration and CMT2A disease. We used the conserved yeast mitofusin FZO1 to study the molecular consequences of CMT2A mutations on Fzo1 function in vivo and in vitro. One mutation (analogous to the CMT2A I213T substitution in the GTPase domain of Mfn2) not only abolishes GTP hydrolysis and mitochondrial membrane fusion but also reduces Mdm30-mediated ubiquitylation and degradation of the mutant protein. Importantly, complexes of wild type and the mutant Fzo1 protein are GTPase active and restore ubiquitylation and degradation of the latter. These studies identify diverse and unexpected effects of CMT2A mutations, including a possible role for mitofusin ubiquitylation and degradation in CMT2A pathogenesis, and provide evidence for a novel link between Fzo1 GTP hydrolysis, ubiquitylation, and mitochondrial fusion.
Rescue of Munc18-1 and -2 Double Knockdown Reveals the Essential Functions of Interaction between Munc18 and Closed Syntaxin in PC12 Cells
Authors: Han Liping , Jiang Tiandan , Han Gayoung A. , Malintan Nancy T. , Xie Li , Wang Li , Tse Frederick W. , Gaisano Herbert Y. , Collins Brett M. , Meunier Frederic A. , Sugita Shuzo
Published: October 2009

Munc18-1 binds to syntaxin-1A via two distinct sites referred to as the “closed” conformation and N terminus binding. The latter has been shown to stimulate soluble N-ethylmaleimide-sensitive factor attachment protein receptor-mediated exocytosis, whereas the former is believed to be inhibitory or dispensable. To precisely define the contributions of each binding mode, we have engineered Munc18-1/-2 double knockdown neurosecretory cells and show that not only syntaxin-1A and -1B but also syntaxin-2 and -3 are significantly reduced as a result of Munc18-1 and -2 knockdown. Syntaxin-1 was mislocalized and the regulated secretion was abolished. We next examined the abilities of Munc18-1 mutants to rescue the defective phenotypes. Mutation (K46E/E59K) of Munc18-1 that selectively prevents binding to closed syntaxin-1 was unable to restore syntaxin-1 expression, localization, or secretion. In contrast, mutations (F115E/E132A) of Munc18-1 that selectively impair binding to the syntaxin-1 N terminus could still rescue the defective phenotypes. Our results indicate that Munc18-1 and -2 act in concert to support the expression of a broad range of syntaxins and to deliver syntaxin-1 to the plasma membrane. Our studies also indicate that the binding to the closed conformation of syntaxin is essential for Munc18-1 stimulatory action, whereas the binding to syntaxin N terminus plays a more limited role in neurosecretory cells.
Efficacy and Safety Assessment of T. Angelica Herbal Tonic, a Phytomedicinal Product Popularly Used in Nigeria
Authors: Esimone Charles O. , Akah Peter A. , Nworu Chukwuemeka S.
Published: October 2009

T. Angelica Herbal Tonic (TAHT) is a herbal product indicated for indigestion and constipation and highly patronized in Nigeria. In this study, the efficacy and safety of the herbal tonic in relation to the label claims were assessed. The effect on peristalsis in mice was evaluated by the charcoal meal model and in vitro using guinea pig ileum. The effects of TAHT on behavior, fertility, birth and organ weights were also determined. Teratogenic potential and reproductive toxicity were studied in pregnant rats. Acute toxicity studies showed that at doses above 5000 mg kg−1, the herbal tonic did not cause lethality and produced no signs of intoxication in mice. The study did not show any gross behavioral changes in mice treated with 1000 mg kg−1 of TAHT as compared with the negative control treatment. TAHT (400 mg kg−1) exhibited a dose‐dependent enhancement in the gastrointestinal tract motility in mice when compared with the negative control. At concentrations up to 300 μg mL−1, TAHT did not cause any significant effect on acetylcholine, histamine and nicotine‐evoked contractions of guinea pig ileum preparation. It took an average of 31.25 ± 4.52 days for the TAHT‐treated animals to litter, which is significantly (P < .05) different from the 55 ± 4.51 days recorded for the control treatment group. TAHT exhibited a modest fertility‐promoting effect and showed lack of abortifacient and teratogenic properties in the study. Generally, the results of this study showed some favorable pharmacological effects of TAHT in animals which may authenticate some of the label claims.
Adjusting for clinical covariates improves the ability of B‐type natriuretic peptide to distinguish cardiac from non‐cardiac dyspnoea: a sub‐study of HEARD‐IT
Authors: Rogers R. Kevin , Stehlik Josef , Stoddard Gregory J. , Greene Thomas , Collins Sean P. , Peacock W. Frank , Maisel Alan D. , Clopton Paul , Michaels Andrew D.
Published: October 2009

Aims

We sought to create a model that adjusts B‐type natriuretic peptide (BNP) for specific covariates to better distinguish cardiac from non‐cardiac dyspnoea.

Methods and results

HEARD‐IT was a multicentre, prospective study of the diagnostic utility of acoustic cardiography in the emergency department. Dyspnoeic patients more than 40 years were eligible. Two cardiologists independently adjudicated the HF outcome. Using logistic regression, a model adjusting BNP for pertinent covariates was developed (n = 740). The mean age was 66 ± 13 years. Age, gender, ethnicity, body mass index, blood urea nitrogen, and creatinine affected BNP levels independently of HF. The model adjusting BNP for these covariates improved the area under receiver operator characteristic curve for HF compared with BNP alone (0.948, 95% CI 0.934–0.963 vs. 0.937, 95% CI 0.920–0.954; P = 0.004). Net reclassification improvement, a novel metric of model performance, was 3.5% for those without HF (P = 0.05) compared with conventional, unadjusted BNP cut‐offs. Thirteen of 116 (11%) patients without HF, but with unadjusted BNP values ≥100 pg/mL, were correctly reclassified as not having HF with the adjusted BNP model.

Conclusion

Adjusting BNP for important covariates may improve its ability to distinguish cardiac from non‐cardiac dyspnoea.
Mathematical models for assessing the role of airflow on the risk of airborne infection in hospital wards
Authors: Noakes Catherine J. , Sleigh P. Andrew
Published: October 2009

Understanding the risk of airborne transmission can provide important information for designing safe healthcare environments with an appropriate level of environmental control for mitigating risks. The most common approach for assessing risk is to use the Wells–Riley equation to relate infectious cases to human and environmental parameters. While it is a simple model that can yield valuable information, the model used as in its original presentation has a number of limitations. This paper reviews recent developments addressing some of the limitations including coupling with epidemic models to evaluate the wider impact of control measures on disease progression, linking with zonal ventilation or computational fluid dynamics simulations to deal with imperfect mixing in real environments and recent work on dose–response modelling to simulate the interaction between pathogens and the host. A stochastic version of the Wells–Riley model is presented that allows consideration of the effects of small populations relevant in healthcare settings and it is demonstrated how this can be linked to a simple zonal ventilation model to simulate the influence of proximity to an infector. The results show how neglecting the stochastic effects present in a real situation could underestimate the risk by 15 per cent or more and that the number and rate of new infections between connected spaces is strongly dependent on the airflow. Results also indicate the potential danger of using fully mixed models for future risk assessments, with quanta values derived from such cases less than half the actual source value.
Effects of microstructures on mesoscopic morphological transitions in deposition growth models
Authors: Sánchez Pedro A. , Sintes Tomás , Piro Oreste , Cartwright Julyan H. E.
Published: October 2009

We study the influence of competing microstructure symmetries on the emergence of different morphologies during the growth by vapour deposition of thin solid films. We perform extensive numerical simulations with a minimal model that includes different microstructures, as well as thermal surface diffusion, to compute the corresponding structure zone model (SZM) and analyse, with statistical physics techniques, the details of transitional morphologies at border zones. We show that the maximum coordination number of the underlying microstructure provides a classification of the statistics of the transitional morphologies at the border between zones I (porous structures) and II (columnar faceted structures) of the SZM.
A generalized theory of viscous and inviscid flutter
Authors: Mandre Shreyas , Mahadevan L.
Published: October 2009

We present a unified theory of flutter in inviscid and viscous flows interacting with flexible structures based on the phenomenon of 1 : 1 resonance. We show this by treating four extreme cases corresponding to viscous and inviscid flows in confined and unconfined flows. To see the common mechanism clearly, we consider the limit when the frequencies of the first few elastic modes are closely clustered and small relative to the convective fluid time scale. This separation of time scales slaves the hydrodynamic force to the instantaneous elastic displacement and allows us to calculate explicitly the dependence of the critical flow speed for flutter on the various problem parameters. We show that the origin of the instability lies in the coincidence of the real frequencies of the first two modes at a critical flow speed beyond which the frequencies become complex, thus making the system unstable to oscillations. This critical flow speed depends on the difference between the frequencies of the first few modes and the nature of the hydrodynamic coupling between them. Our generalized framework applies to a range of elastohydrodynamic systems and further extends the Benjamin–Landahl classification of fluid–elastic instabilities.
Recent advances in Chinese palaeontology
Authors: Xu Xing , Luo Zhe-Xi , Rong Jia-Yu
Published: October 2009

Discoveries are a driving force for progress in palaeontology. Palaeontology as a discipline of scientific inquiry has gained many fresh insights into the history of life, from the discoveries of many new fossils in China in the last 20 years, and from the new ideas derived from these fossils. This special issue of
Proceedings of Royal Society B
entitled
Recent Advances in Chinese Palaeontology
selects some of the very latest studies aimed at resolving the current problems of palaeontology and evolutionary biology based on new fossils from China. These fossils and their studies help to clarify some historical debates about a particular fossil group, or to raise new questions about history of life, or to pose a new challenge in our pursuit of science. These works on new Chinese fossils have covered the whole range of the diversity through the entire Phanerozoic fossil record.
Replication Protein A (RPA1a) Is Required for Meiotic and Somatic DNA Repair But Is Dispensable for DNA Replication and Homologous Recombination in Rice
Authors: Chang Yuxiao , Gong Liang , Yuan Wenya , Li Xingwang , Chen Guoxing , Li Xianghua , Zhang Qifa , Wu Changyin
Published: October 2009

Replication protein A (RPA), a highly conserved single-stranded DNA-binding protein in eukaryotes, is a stable complex comprising three subunits termed RPA1, RPA2, and RPA3. RPA is required for multiple processes in DNA metabolism such as replication, repair, and homologous recombination in yeast (Saccharomyces cerevisiae) and human. Most eukaryotic organisms, including fungi, insects, and vertebrates, have only a single RPA gene that encodes each RPA subunit. Arabidopsis (Arabidopsis thaliana) and rice (Oryza sativa), however, possess multiple copies of an RPA gene. Rice has three paralogs each of RPA1 and RPA2, and one for RPA3. Previous studies have established their biochemical interactions in vitro and in vivo, but little is known about their exact function in rice. We examined the function of OsRPA1a in rice using a T-DNA insertional mutant. The osrpa1a mutants had a normal phenotype during vegetative growth but were sterile at the reproductive stage. Cytological examination confirmed that no embryo sac formed in female meiocytes and that abnormal chromosomal fragmentation occurred in male meiocytes after anaphase I. Compared with wild type, the osrpa1a mutant showed no visible defects in mitosis and chromosome pairing and synapsis during meiosis. In addition, the osrpa1a mutant was hypersensitive to ultraviolet-C irradiation and the DNA-damaging agents mitomycin C and methyl methanesulfonate. Thus, our data suggest that OsRPA1a plays an essential role in DNA repair but may not participate in, or at least is dispensable for, DNA replication and homologous recombination in rice.
Flexible Tools for Gene Expression and Silencing in Tomato
Authors: Fernandez Ana I. , Viron Nicolas , Alhagdow Moftah , Karimi Mansour , Jones Matthew , Amsellem Ziva , Sicard Adrien , Czerednik Anna , Angenent Gerco , Grierson Donald , May Sean , Seymour Graham , Eshed Yuval , Lemaire-Chamley Martine , Rothan Christophe , Hilson Pierre
Published: October 2009

As a genetic platform, tomato (Solanum lycopersicum) benefits from rich germplasm collections and ease of cultivation and transformation that enable the analysis of biological processes impossible to investigate in other model species. To facilitate the assembly of an open genetic toolbox designed to study Solanaceae, we initiated a joint collection of publicly available gene manipulation tools. We focused on the characterization of promoters expressed at defined time windows during fruit development, for the regulated expression or silencing of genes of interest. Five promoter sequences were captured as entry clones compatible with the versatile MultiSite Gateway format: PPC2, PG, TPRP, and IMA from tomato and CRC from Arabidopsis (Arabidopsis thaliana). Corresponding transcriptional fusions were made with the GUS gene, a nuclear-localized GUS-GFP reporter, and the chimeric LhG4 transcription factor. The activity of the promoters during fruit development and in fruit tissues was confirmed in transgenic tomato lines. Novel Gateway destination vectors were generated for the transcription of artificial microRNA (amiRNA) precursors and hairpin RNAs under the control of these promoters, with schemes only involving Gateway BP and LR Clonase reactions. Efficient silencing of the endogenous phytoene desaturase gene was demonstrated in transgenic tomato lines producing a matching amiRNA under the cauliflower mosaic virus 35S or PPC2 promoter. Lastly, taking advantage of the pOP/LhG4 two-component system, we found that well-characterized flower-specific Arabidopsis promoters drive the expression of reporters in patterns generally compatible with heterologous expression. Tomato lines and plasmids will be distributed through a new Nottingham Arabidopsis Stock Centre service unit dedicated to Solanaceae resources.
A Cytoplasmically Inherited Barley Mutant Is Defective in Photosystem I Assembly Due to a Temperature-Sensitive Defect in ycf3 Splicing
Authors: Landau Alejandra Mabel , Lokstein Heiko , Scheller Henrik Vibe , Lainez Verónica , Maldonado Sara , Prina Alberto Raúl
Published: October 2009

A cytoplasmically inherited chlorophyll-deficient mutant of barley (Hordeum vulgare) termed cytoplasmic line 3 (CL3), displaying a viridis (homogeneously light-green colored) phenotype, has been previously shown to be affected by elevated temperatures. In this article, biochemical, biophysical, and molecular approaches were used to study the CL3 mutant under different temperature and light conditions. The results lead to the conclusion that an impaired assembly of photosystem I (PSI) under higher temperatures and certain light conditions is the primary cause of the CL3 phenotype. Compromised splicing of ycf3 transcripts, particularly at elevated temperature, resulting from a mutation in a noncoding region (intron 1) in the mutant ycf3 gene results in a defective synthesis of Ycf3, which is a chaperone involved in PSI assembly. The defective PSI assembly causes severe photoinhibition and degradation of PSII.
Systematic review: distribution of advanced neoplasia according to polyp size at screening colonoscopy
Authors: HASSAN C. , PICKHARDT P. J. , KIM D. H. , DI GIULIO E. , ZULLO A. , LAGHI A. , REPICI A. , IAFRATE F. , OSBORN J. , ANNIBALE B.
Published: October 2009

Background  The impact of not referring sub‐centimetre polyps identified at CT colonography upon the efficacy of colorectal cancer screening remains uncertain.

Aim  To determine the distribution of advanced neoplasia according to polyp size in a screening setting.

Methods  Published studies reporting the distribution of advanced adenomas in asymptomatic screening cohorts according to polyp size were identified by MEDLINE and EMBASE searches. Predefined outputs were the screening rates of advanced adenomas represented by diminutive (≦5 mm), small (6–9 mm), sub‐centimetre (<10 mm) and large (≧10 mm) polyp sizes.

Results  Data from four studies with 20 562 screening subjects met the primary inclusion criteria. Advanced adenomas were detected in 1155 (5.6%) subjects (95% CI = 5.3–5.9), corresponding to diminutive, small and large polyps in 4.6% (95% CI = 3.4–5.8), 7.9% (95% CI = 6.3–9.4) and 87.5% (95% CI = 86–89.4) of cases respectively. The frequency of advanced lesions among patients whose largest polyp was diminutive, small, sub‐centimetre and large in size was 0.9%, 4.9%, 1.7% and 73.5% respectively.

Conclusions  Based on this systematic review, a 6‐mm polyp size threshold for polypectomy referral would identify over 95% of subjects with advanced adenomas, whereas a 10‐mm threshold would identify 88% of cases.

Aliment Pharmacol Ther 31, 210–217
Methylenetetrahydrofolate reductase (MTHFR) gene polymorphisms and FOLFOX response in colorectal cancer patients
Authors: Etienne‐Grimaldi Marie‐Christine , Milano Gérard , Maindrault‐Gœbel Frédérique , Chibaudel Benoist , Formento Jean‐Louis , Francoual Mireille , Lledo Gérard , André Thierry , Mabro May , Mineur Laurent , Flesch Michel , Carola Elisabeth , De Gramont Aimery
Published: October 2009

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT

• Numerous clinical studies, including a few prospective ones, have reported conflicting results on the impact of gene polymorphisms related to fluorouracil (FU) and oxaliplatin pharmacodynamics.

WHAT THIS STUDY ADDS

• This prospective study is the first to report that clinical response to FOLFOX is significantly related to methylenetetrahydrofolate reductase (MTHFR) gene polymorphisms (677C→T and 1298A→C), with a response rate of 37, 53, 63 and 80% in patients harbouring no, one, two or three favourable MTHFR alleles, respectively.

• Only polymorphisms of genes related to oxaliplatin pharmacodynamics (GSTπ 105Ile→Val and XPD 751Ly→Gln) influenced progression‐free survival.

• These results corroborate the observation that response was related to the cumulative FU dose, whereas progression‐free survival was related to the cumulative oxaliplatin dose.

AIMS To test prospectively the predictive value of germinal gene polymorphisms related to fluorouracil (FU) and oxaliplatin (Oxa) pharmacodynamics on toxicity and responsiveness of colorectal cancer (CRC) patients receiving FOLFOX therapy.

METHODS Advanced CRC patients (n= 117) receiving FOLFOX 7 therapy were enrolled. Gene polymorphisms relevant for FU [thymidylate synthase (TYMS, 28 bp repeats including the G→C mutation + 6 bp deletion in 3'UTR), methylenetetrahydrofolate reductase (MTHFR, 677C→T, 1298A→C), dihydropyrimidine deshydrogenase (IVS14+1G→A) and Oxa: glutathione S‐transferase (GST) π (105Ile→Val, 114Ala→Val), excision repair cross‐complementing group 1 (ERCC1) (118AAT→AAC), ERCC2 (XPD, 751Lys→Gln) and XRCC1 (399Arg→Gln)] were determined (blood mononuclear cells).

RESULTS None of the genotypes was predictive of toxicity. Response rate (54.7% complete response + partial response) was related to FU pharmacogenetics, with both 677C→T (P= 0.042) and 1298A→C (P= 0.004) MTHFR genotypes linked to clinical response. Importantly, the score of favourable MTHFR alleles (677T and 1298C) was positively linked to response, with response rates of 37.1, 53.3, 62.5 and 80.0% in patients bearing no, one, two or three favourable alleles, respectively (P= 0.040). Polymorphisms of genes related to Oxa pharmacodynamics showed an influence on progression‐free survival, with a better outcome in patients bearing GSTπ 105 Val/Val genotype or XPD 751Lys‐containing genotype (P= 0.054).

CONCLUSIONS These results show that response to FOLFOX therapy in CRC patients may be driven by MTHFR germinal polymorphisms.
The fundamental contribution of William Bate Hardy to shape the concept of mast cell heterogeneity
Authors: Crivellato Enrico , Ribatti Domenico
Published: October 2009

This review article acknowledges the pioneering contribution of William Bate Hardy in shaping the concept of mast cell heterogeneity. In two outstanding papers, published in 1894 and 1895, he focussed on the ‘wandering cells’ (the modern leucocytes) in different mammalian species and distinguished two types of granular basophil cells, i.e., the coarsely granular basophil cells and the splanchnic basophil cells. These corresponded to the populations of connective tissue‐type and mucosal mast cells, respectively, described 70 years later by Enerbäck in rodents. Among the coarsely granular basophil cells, he also differentiated those cells which populated the serosal cavities – the so‐called coelomic coarsely granular basophil cells – from the common coarsely granular basophil cells, which were localized in the connective tissues. He stated that the granular basophil cells presented with different morphological and histochemical characteristics in diverse animal species as well as at different anatomical sites. Remarkably, he performed a series of functional experiments on the basophil cells as well as the other wandering cells, and suggested the view that different granular basophil cells might express functional specializations.
Lactic acid bacteria differ in their ability to induce functional regulatory T cells in humans
Authors: De Roock S. , Van Elk M. , Van Dijk M. E. A. , Timmerman H. M. , Rijkers G. T. , Prakken B. J. , Hoekstra M. O. , De Kleer I. M.
Published: October 2009

Background
Trials with probiotic lactic acid bacteria have yielded different results, which may be due to the strains used. Lactobacilli and bifidobacteria are known to be potent modulators of the immune system. The capacity of these bacteria used as probiotics to influence both T helper type 1 (Th1)‐ and Th2‐mediated diseases has been shown before. However, the ability of strains to induce forkhead box P3 (FOXP3+) expressing regulatory T cells has not yet been investigated.

Objective
Test the inherent differences between strains in their capacity to induce functional regulatory T cells in human peripheral blood mononuclear cells (PBMC).

Methods
Human PBMC were co‐cultured in vitro with either Bifidobacterium lactis W51, Lactobacillus acidophilus W55 or Lactobacillus plantarum W62 or an Escherichia coli control strain. The percentage of FOXP3+ cells, the origin of the induced cells and the functionality of these cells were assessed.

Results
Probiotic strains differ in their capacity to induce regulatory T cells. FOXP3+ cells were induced from CD25− cells and were able to suppress effector T cells. Naturally occurring regulatory T cells were not affected by co‐culture with lactobacilli. IL‐10 concentrations found in the supernatant showed a trend towards the same differences between strains. Blockade of IL‐10 did not influence the up‐regulation of FOXP3. No differences between lactic acid bacteria were found in IL‐17, IFN‐γ or IL‐13.

Conclusions
Some probiotic strains are potent inducers of regulatory cells, while others are not. The clear differences between strains imply that an in vitro characterization of probiotic strains before application is recommended.

Cite this as: S. de Roock, M. van Elk, M. E. A. van Dijk, H. M. Timmerman, G. T. Rijkers, B. J. Prakken, M. O. Hoekstra and I. M. de Kleer, Clinical & Experimental Allergy, 2010 (40) 103–110.
The effect of oral steroids with and without vitamin D3 on early efficacy of immunotherapy in asthmatic children
Authors: Majak P. , Rychlik B. , Stelmach I.
Published: October 2009

Background
The possibility of additional strategies to enhance the effectiveness of specific immunotherapy (SIT) is highly attractive.

Aim
The aim of our study was to assess the influence of oral corticosteroids and oral corticosteroids combined with vitamin D3 on the early clinical and immunological effects of SIT.

Methods
It was a randomized, double‐blind, placebo‐controlled trial conducted in 54 asthmatic children allergic to house dust mites. Intervention was based on receiving a single dose of oral steroid, with or without vitamin D3, or placebo only on the day of the build‐up phase of SIT.

Results
After 12 months of SIT, the median daily inhaled corticosteroid (ICS) dose, which controls the symptoms of asthma, was reduced by 25% in the steroid group. However, a 50% reduction of the median daily ICS dose was observed in the control group. The clinical effects of SIT were not affected in the steroid+D3 group. Concomitantly, we found that intervention with prednisone significantly impaired the induction of T regulatory lymphocytes. Importantly, the clinical and immunological effects of SIT were not affected by intervention with steroids administered with vitamin D3.

Conclusions
Our study failed to show a beneficial effect of oral corticosteroids on allergen‐specific immunotherapy. We observed that the combined administration of a corticosteroid drug and allergen extract suppressed the early clinical and immunological effects of SIT and that vitamin D3 prevented this ‘adverse’ influence of steroids.
Suppression of gastric acid increases the risk of developing Immunoglobulin E‐mediated drug hypersensitivity: human diclofenac sensitization and a murine sensitization model
Authors: Riemer A. B. , Gruber S. , Pali‐Schöll I. , Kinaciyan T. , Untersmayr E. , Jensen‐Jarolim E.
Published: October 2009

Background
Hypersensitivity reactions towards non‐steroidal anti‐inflammatory drugs (NSAID) are common, although true allergies are detectable only in a subgroup of patients. The current study was prompted by a case observation, where a patient experienced generalized urticaria following his second course of diclofenac and proton pump inhibitor medication, and was found to have diclofenac‐specific IgE. During recent years, our group has been investigating the importance of gastric digestion in the development of food allergies, demonstrating anti‐acid medication as a risk factor for sensitization against food proteins.

Objective
Here, we aimed to investigate whether the mechanism of food allergy induction described can also be causative in NSAID allergy, using diclofenac as a paradigm.

Methods
We subjected BALB/c mice to several oral immunization regimens modelled after the patient's medication intake. Diclofenac was applied with or without gastric acid suppression, in various doses, alone or covalently coupled to albumin, a protein abundant in gastric juices. Immune responses were assessed on the antibody level, and functionally examined by in vitro and in vivo crosslinking assays.

Results
Only mice receiving albumin‐coupled diclofenac under gastric acid suppression developed anti‐diclofenac IgG1 and IgE, whereas no immune responses were induced by the drug alone or without gastric acid suppression. Antibody induction was dose dependent with the group receiving the higher dose of the drug showing sustained anti‐diclofenac titres. The antibodies induced triggered basophil degranulation in vitro and positive skin tests in vivo.

Conclusion
Gastric acid suppression was found to be a causative mechanism in the induction of IgE‐mediated diclofenac allergy.

Cite this as: A. B. Riemer, S. Gruber, I. Pali‐Schöll, T. Kinaciyan, E. Untersmayr and E. Jensen‐Jarolim, Clinical & Experimental Allergy, 2010 (40) 486–493.
The effect of ultrasonic removal of various root‐end filling materials
Authors: Pirani C. , Iacono F. , Chersoni S. , Sword J. , Pashley D. H. , Tay F. R. , Looney S. , Gandolfi M. G. , Prati C.
Published: October 2009

Aim  To compare residual root‐end filling material in apical root‐end cavities following their removal with ultrasonic retrotips.

Methodology  Thirty single‐rooted teeth were filled with Thermafil and AH Plus sealer. Root‐ends were resected at 90°, 3 mm from the apex. Root‐end cavities were prepared with diamond burs and ultrasonic retrotips and filled with one of three filling materials: group I: Retro‐TC (calcium silicate‐based cement), group II: IRM (Dentsply, Germany), group III: Vitrebond (3M ESPE, USA). After 30 days of storage, ultrasonic retrotips were used to remove materials from the root‐end cavities. The ultrasonic application time was fixed at 60 s. Polyether impressions and replicas of the root‐ends were made. Root apices and replicas were examined by one operator under a scanning electron microscope. Remnants of residual materials were evaluated using a four‐level scoring system; fractures, smear layer and exposed dentinal tubules were also examined.

Results  Forty per cent of the specimens filled with Retro‐TC revealed complete removal of the material with exposure of dentinal tubules, whilst 60% contained residual cement. Twenty per cent of specimens filled with IRM were completely devoid of material, whereas 80% had retained material. Ten per cent of specimens filled with Vitrebond retained a moderate amount of material whilst 90% had substantial retention of the material. Statistically significant differences were found (P < 0.05) amongst the three groups of materials.

Conclusions  Retro‐TC was successfully removed in 40% of cases using ultrasonics retrotips for 60 s, whereas IRM and Vitrebond specimens had evidence of retained material in 80% and 90% of all specimens respectively.
An in vivo comparison of working length determination of two frequency‐based electronic apex locators
Authors: Pascon E. A. , Marrelli M. , Congi O. , Ciancio R. , Miceli F. , Versiani M. A.
Published: October 2009

Aim  To compare in vivo the accuracy of two electronic apex locators (EALs) by means of digital radiographic imaging system.

Methodology  Electronic working lengths of 831 canals were determined with the DentaPort ZX and Raypex 5 apex locators and confirmed radiographically. The radiographic images acquired with the aid of a digital radiographic imaging system (VisualiX eHD; Gendex Dental Systems, Des Plaines, IL, USA) were blindly analysed by two independent evaluators. The distance between the file tip and the radiographic apex was measured using dedicated software (VixWin Pro, Gendex Dental Systems, Des Plaines, IL, USA) and the mean distance achieved between different tooth type and EALs were compared statistically. Statistical analyses were performed using the t‐test for independent samples and one‐way anova with the null hypothesis set as 5%. Positive or negative values were recorded when the file tip was detected beyond or short of the radiographic apex, respectively.

Results  The mean distance between file tip and radiographic apex were −1.08 ± 0.73 and −1.0 ± 0.67 mm considering DentaPort ZX and Raypex 5 groups, respectively, with no significant differences (P > 0.05). No statistically significant differences were found amongst the same tooth type when comparing both groups (P > 0.05) or amongst different teeth type in the same group (P > 0.05).

Conclusions  Within the limitations of this in vivo study, the DentaPort ZX and Raypex 5 were similar in terms of accuracy.
Identification of a novel FAM83H mutation and microhardness of an affected molar in autosomal dominant hypocalcified amelogenesis imperfecta
Authors: Hyun H.‐K. , Lee S.‐K. , Lee K.‐E. , Kang H.‐Y. , Kim E.‐J. , Choung P.‐H. , Kim J.‐W.
Published: October 2009

Aim  To determine the underlying molecular genetic aetiology of a family with the hypocalcified form of amelogenesis imperfecta and to investigate the hardness of the enamel and dentine of a known FAM83H mutation.

Methodology  Mutational screening of the FAM83H on the basis of candidate gene approach was performed. All exons and exon–intron boundaries was amplified and sequenced. A microhardness test was performed to measure the Vickers microhardness value.

Results  A novel nonsense mutation (c.1354C>T, p.Q452X) was identified in the last exon of FAM83H, which resulted in soft, uncalcified enamel. The affected enamel was extremely soft (about 17% of the normal control), but the underlying dentine was as hard as the normal control.

Conclusions  Mutational analysis revealed a novel mutation in FAM83H gene. Hardness of dentine was not affected by the mutation, whilst the enamel was extremely soft.
Investigation of the root canal configuration of mandibular first molars in a Taiwan Chinese population
Authors: Chen G. , Yao H. , Tong C.
Published: October 2009

Aim  To investigate the root form and canal morphology of mandibular first molar teeth in a Taiwan Chinese population.

Methodology  A total of 183 mandibular first molars were collected and examined visually through a dissecting microscope after rendering the teeth transparent. The number of roots, number of canals and canal configuration were recorded. The root canal morphology was categorized and compared according to Vertucci (1984).

Results  Overall 46% of the mandibular first molars had four canals and 20% had extra‐distal roots (distolingual root). In addition, 97% of the specimens had two mesial canals and 46% had two distal canals. In total, 68% of teeth with two mesial canals had two separated apical foramina, and 33% of teeth with two distal canals had two apical foramina. The most common canal configurations (Vertucci, 1984) of mesial roots were type 4 (46%) and type 2 (23%). Type 1 was encountered in 54% of the distal roots.

Conclusions  The frequency of the extra‐distal root on the mandibular first molar was 20%, and the incidence of three canals (Vertucci’s type 8 classification) with separate apical foramina at the apex of the mesial root was 6%. The three‐rooted variation of the mandibular first molar appears to be a genetic characteristic of an Asiatic racial background.
Bridging the gap between community ecology and historical biogeography: niche conservatism and community structure in emydid turtles
Authors: STEPHENS P. R. , WIENS J. J.
Published: October 2009

Historical (phylogenetic) biogeography and community ecology were once integrated as part of the broader study of organismal diversity, but in recent decades have become largely separate disciplines. This is unfortunate because many patterns studied by community ecologists may originate through processes studied by historical biogeographers and vice versa. In this study, we explore the causes of a geographic pattern of community structure (habitat use) in the emydid turtle assemblages of eastern North America, with more semi‐terrestrial species of the subfamily Emydinae in the north and more aquatic species of Deirochelyinae in the south. Specifically, we address the factors that prevent northern emydines from invading southern communities. We test for competitive exclusion by examining patterns of range overlap, and test for the role of niche conservatism using analyses of climatic and physiological data based on a multilocus molecular phylogeny. We find no support for competitive exclusion, whereas several lines of evidence support the idea that niche conservatism has prevented northern emydines from dispersing into southern communities. Our results show how understanding the causes of patterns of historical biogeography may help explain patterns of community structure.
Bacterial chemoattraction towards jasmonate plays a role in the entry of Dickeya dadantii through wounded tissues
Authors: Antunez‐Lamas Maria , Cabrera Ezequiel , Lopez‐Solanilla Emilia , Solano Roberto , González‐Melendi Pablo , Chico Jose Manuel , Toth Ian , Birch Paul , Pritchard Leighton , Liu Hui , Rodriguez‐Palenzuela Pablo
Published: October 2009

Jasmonate is a key signalling compound in plant defence that is synthesized in wounded tissues. In this work, we have found that this molecule is also a strong chemoattractant for the phythopathogenic bacteria Dickeya dadantii (ex‐Erwinia chysanthemi). Jasmonic acid induced the expression of a subset of bacterial genes possibly involved in virulence/survival in the plant apoplast and bacterial cells pre‐treated with jasmonate showed increased virulence in chicory and Saintpaulia leaves. We also showed that tissue wounding induced bacterial spread through the leaf surface. Moreover, the jasmonate‐deficient aos1 Arabidopsis thaliana mutant was more resistant to bacterial invasion by D. dadantii than wild‐type plants. These results are consistent with the hypothesis that sensing jasmonic acid by this bacterium helps the pathogen to ingress inside plant tissues.
SpoIIIE and a novel type of DNA translocase, SftA, couple chromosome segregation with cell division in Bacillus subtilis
Authors: Kaimer Christine , González‐Pastor José Eduardo , Graumann Peter L.
Published: October 2009

Cell division must only occur once daughter chromosomes have been fully separated. However, the initiating event of bacterial cell division, assembly of the FtsZ ring, occurs while chromosome segregation is still ongoing. We show that a two‐step DNA translocase system exists in Bacillus subtilis that couples chromosome segregation and cell division. The membrane‐bound DNA translocase SpoIIIE assembled very late at the division septum, and only upon entrapment of DNA, while its orthologue, SftA (YtpST), assembled at each septum in B. subtilis soon after FtsZ. Lack of SftA resulted in a moderate segregation defect at a late stage in the cell cycle. Like the loss of SpoIIIE, the absence of SftA was deleterious for the cells during conditions of defective chromosome segregation, or after induction of DNA damage. Lack of both proteins exacerbated all phenotypes. SftA forms soluble hexamers in solution, binds to DNA and has DNA‐dependent ATPase activity, which is essential for its function in vivo. Our data suggest that SftA aids in moving DNA away from the closing septum, while SpoIIIE translocates septum‐entrapped DNA only when septum closure precedes complete segregation of chromosomes.