bluuebunny/crossref_metadata_2025_split huggingface dataset:
Genetic Analysis of Transfer-Related Regions of the Vancomycin Resistance
Enterococcus
Conjugative Plasmid pHTβ: Identification of
oriT
and a Putative Relaxase Gene
Authors: Tomita Haruyoshi , Ike Yasuyoshi 
Published: November 2005

ABSTRACT

The pHT plasmids pHTα (65.9 kbp), pHTβ (63.7 kbp), and pHTγ (66.5 kbp) are highly conjugative pheromone-independent pMG1-like plasmids that carry Tn
1546
-like transposons encoding vancomycin resistance. pHTβ is the prototype plasmid, and the pHTα and pHTγ plasmids are derivatives of the insertion into pHTβ of an IS
232
-like (2.2 kbp) element and a group II intron (2.8 kbp), respectively. The complete nucleotide sequence of the pHTβ plasmid was determined and, with the exception of the Tn
1546
-like insertion (10,851 bp), was found to be 52,890 bp. Sixty-one open reading frames (ORFs) having the same transcript orientation were identified. A homology search revealed that 22 of the pHTβ (pHT) plasmid ORFs showed similarities to the ORFs identified on the pXO2 plasmid (96.2 kbp), which is the virulence plasmid essential for capsule formation by
Bacillus anthracis
; however, the functions of most of the ORFs remain unknown. Most other ORFs did not show any significant homology to reported genes for which functions have been analyzed. To investigate the highly efficient transfer mechanism of the pHT plasmid, mutations with 174 unique insertions of transposon Tn
917-lac
insertion mutants of pHTβ were obtained. Of the 174 derivatives, 92 showed decrease or loss in transfer frequency, and 74 showed normal transfer frequency and LacZ expression. Eight derivatives showed normal transfer and no LacZ expression. Inserts within the 174 derivatives were mapped to 124 different sites on pHTβ. The Tn
917-lac
insertions which resulted in altered transfer frequency mapped to three separate regions designated I, II, and III, which were separated by segments in which insertions of Tn
917
-
lac
did not affect transfer. There was no region homologous to the previously reported
oriT
sequences in the pHT plasmid. The
oriT
was cloned by selection for the ability to mobilize the vector plasmid pAM401. The
oriT
region resided in a noncoding region (192 bp) between ORF31 and ORF32 and contained three direct repeat sequences and two inverted repeat sequences. ORF34, encoding a 506-amino-acid protein which was located downstream of the
oriT
region, contains the three conserved motifs (I to III) of the DNA relaxase/nickase of mobile plasmids. The transfer abilities of the Tn
917-lac
-insertion mutants of ORF34 or a mutant of ORF34 with an in-frame motif III deletion were completely abolished. The sequence of the
oriT
region and the deduced relaxase/nickase protein of ORF34 showed no significant similarity to the
oriT
and relaxase/nickase of other conjugative plasmids, respectively. The putative relaxase/nickase protein of ORF34 could be classified as a new member of the MOB
MG
family.
Functional Genomic Analysis of Three Nitrogenase Isozymes in the Photosynthetic Bacterium
Rhodopseudomonas palustris
Authors: Oda Yasuhiro , Samanta Sudip K. , Rey Federico E. , Wu Liyou , Liu Xiudan , Yan Tingfen , Zhou Jizhong , Harwood Caroline S. 
Published: November 2005

ABSTRACT

The photosynthetic bacterium
Rhodopseudomonas palustris
is one of just a few prokaryotes described so far that has
vnf
and
anf
genes for alternative vanadium cofactor (V) and iron cofactor (Fe) nitrogenases in addition to
nif
genes for a molybdenum cofactor (Mo) nitrogenase. Transcriptome data indicated that the 32 genes in the
nif
gene cluster, but not the
anf
or
vnf
genes, were induced in wild-type and Mo nitrogenase-expressing strains grown under nitrogen-fixing conditions in Mo-containing medium. Strains that were unable to express a functional Mo nitrogenase due to mutations in Mo nitrogenase structural genes synthesized functional V and Fe nitrogenases and expressed
vnf
and
anf
genes in nitrogen-fixing growth media that contained Mo and V at concentrations far in excess of those that repress alternative nitrogenase gene expression in other bacteria. Thus, not only does
R. palustris
have multiple enzymatic options for nitrogen fixation, but in contrast to reports on other nitrogen-fixing bacteria, the expression of its alternative nitrogenases is not repressed by transition metals. Between 95 and 295 genes that are not directly associated with nitrogenase synthesis and assembly were induced under nitrogen-fixing conditions, depending on which nitrogenase was being used by
R. palustris
. Genes for nitrogen acquisition were expressed at particularly high levels during alternative nitrogenase-dependent growth. This suggests that alternative nitrogenase-expressing cells are relatively starved for nitrogen and raises the possibility that fixed nitrogen availability may be the primary signal that controls the synthesis of the V and Fe nitrogenases.
CadC Activates pH-Dependent Expression of the
Vibrio vulnificus cadBA
Operon at a Distance through Direct Binding to an Upstream Region
Authors: Rhee Jee Eun , Kim Kun-Soo , Choi Sang Ho 
Published: November 2005

ABSTRACT

The
Vibrio vulnificus cadBA
genes were transcribed as a transcriptional operon by a single promoter, P

cadBA

, which was activated by CadC in a pH-dependent manner. A direct interaction between CadC and the P

cadBA

DNA was demonstrated, and a CadC binding site centered at −233.5 was mapped by deletion analyses of P

cadBA

and confirmed by a DNase I protection assay.
Cinical and pathological characteristics of Chinese patients with antineutrophil cytoplasmic autoantibody associated systemic vasculitides: a study of 426 patients from a single centre
Authors: Chen M , Yu F , Zhang Y , Zhao M H 
Published: November 2005

AbstractBackgroundAntineutrophil cytoplasmic autoantibodies (ANCA) are serological markers of ANCA associated systemic vasculitides (AASV), which is one of the most common multisystem autoimmune diseases. Features of Chinese patients with AASV have not been fully investigated.ObjectiveTo analyse the clinical and pathological characteristics of Chinese patients with AASV.Methods426 Chinese patients with AASV diagnosed in the past eight years were retrospectively studied and their clinical and pathological data were analysed.ResultsOf the 426 patients, 87 (20.4%) were Wegener’s granulomatosis, 337 (79.1%) were microscopic polyangiitis and two (0.5%) were Churg-Strauss syndrome. Only 201 of 426 (47.2%) patients were diagnosed within three months. Clinically, the patients had multisystem involvement. Altogether 371 of 426 (87.1%) had kidney involvement and 260 of 426 (61.0%) had lung involvement. The prevalences of renal involvement and fatigue were significantly higher in patients with MPO-ANCA than that in patients with PR3-ANCA; the prevalences of ophthalmic, nasal involvement, rash, and arthragia were significantly higher in patients with PR3-ANCA than those in patients with MPO-ANCA. The one and five year death rates were 13.1% and 22.4%, respectively. The percentage of patients progressing to end stage renal disease at one and five years was 15.9% and 27.1%, respectively.ConclusionsAASV is not a rare autoimmune disease in Chinese people. Kidney and lung were the most vulnerable organs. For patients with multiorgan damage, an ANCA test should be performed to make an early diagnosis and to start treatment in time.
Assessment of Low Linear Energy Transfer Radiation–Induced Bystander Mutagenesis in a Three-Dimensional Culture Model
Authors: Persaud Rudranath , Zhou Hongning , Baker Sarah E. , Hei Tom K. , Hall Eric J. 
Published: November 2005

Abstract
A three-dimensional cell culture model composed of human-hamster hybrid (AL) and Chinese hamster ovary (CHO) cells in multicellular clusters was used to investigate low linear energy transfer (LET) radiation–induced bystander genotoxicity. CHO cells were mixed with AL cells in a 1:5 ratio and briefly centrifuged to produce a spheroid of 4 × 106 cells. CHO cells were labeled with tritiated thymidine ([3H]dTTP) for 12 hours and subsequently incubated with AL cells for 24 hours at 11°C. The short-range β-particles emitted by [3H]dTTP result in self-irradiation of labeled CHO cells; thus, biological effects on neighboring AL cells can be attributed to the bystander response. Nonlabeled bystander AL cells were isolated from among labeled CHO cells by using a magnetic separation technique. Treatment of CHO cells with 100 μCi [3H]dTTP resulted in a 14-fold increase in bystander mutation incidence among neighboring AL cells compared with controls. Multiplex PCR analysis revealed the types of mutants to be significantly different from those of spontaneous origin. The free radical scavenger DMSO or the gap junction inhibitor Lindane within the clusters significantly reduced the mutation incidence. The use of AL cells that are dominant negative for connexin 43 and lack gap junction formation produced a complete attenuation of the bystander mutagenic response. These data provide evidence that low LET radiation can induce bystander mutagenesis in a three-dimensional model and that reactive oxygen species and intercellular communication may have a modulating role. The results of this study will address the relevant issues of actual target size and radiation quality and are likely to have a significant effect on our current understanding of radiation risk assessment.
Induction of Early Post-Transplant Graft-versus-Leukemia Effects Using Intentionally Mismatched Donor Lymphocytes and Elimination of Alloantigen-Primed Donor Lymphocytes for Prevention of Graft-versus-Host Disease
Authors: Yung Iris , Weiss Lola , Abdul-Hai Ali , Kasir Judith , Reich Shoshana , Slavin Shimon 
Published: November 2005

AbstractGraft-versus-leukemia (GVL) effects can be induced in tolerant mixed chimeras prepared with nonmyeloablative conditioning. GVL effects can be amplified by post-grafting donor lymphocyte infusion (DLI). Unfortunately, DLI is frequently associated with graft-versus-host disease (GVHD). We investigated the feasibility of induction of potent GVL effects by DLI using intentionally mismatched lymphocytes followed by elimination of alloreactive donor T cells by cyclophosphamide for prevention of lethal GVHD following induction of very short yet most potent GVL effects. Mice inoculated with B-cell leukemia (BCL1) and mismatched donor lymphocytes were treated 2 weeks later with low-dose or high-dose cyclophosphamide. All mice receiving cyclophosphamide 2 weeks after DLI survived GVHD, and no residual disease was detected by PCR; all control mice receiving DLI alone died of GVHD. Analysis of host (female) and donor (male) DNA showed that cyclophosphamide treatment eradicated most alloreactive donor cells, yet mixed chimerism was converted to full donor chimerism following transient self-limited GVHD. Our working hypothesis suggests that short-term yet effective and safe adoptive immunotherapy of leukemia may be accomplished early post-transplantation using alloreactive donor lymphocytes, with prevention of GVHD by elimination of GVL effector cells.
Mammalian Target of Rapamycin, a Molecular Target in Squamous Cell Carcinomas of the Head and Neck
Authors: Amornphimoltham Panomwat , Patel Vyomesh , Sodhi Akrit , Nikitakis Nikolaos G. , Sauk John J. , Sausville Edward A. , Molinolo Alfredo A. , Gutkind J. Silvio 
Published: November 2005

Abstract
Emerging knowledge on how the dysregulated function of signaling networks contributes to the malignant growth of squamous cell carcinoma of the head and neck (HNSCC) can now be exploited to identify novel mechanism-based anticancer treatments. In this regard, we have observed that persistent activation of the serine/threonine kinase Akt is a frequent event in HNSCC, and that blockade of its upstream kinase, 3′-phosphoinositide-dependent kinase 1, potently inhibits tumor cell growth. Akt promotes cell proliferation by its ability to coordinate mitogenic signaling with energy- and nutrient-sensing pathways that control protein synthesis through the atypical serine/threonine kinase, mammalian target of rapamycin (mTOR). This kinase, in turn, phosphorylates key eukaryotic translation regulators, including p70-S6 kinase and the eukaryotic translation initiation factor, 4E binding protein 1. Indeed, we show here that aberrant accumulation of the phosphorylated active form of S6, the most downstream target of the Akt-mTOR-p70-S6 kinase pathway, is a frequent event in clinical specimens from patients with HNSCC and their derived cell lines. Of interest, this enhanced level of the phosphorylated active form of S6 was rapidly reduced in HNSCC cell lines and HNSCC xenograft models at clinically relevant doses of rapamycin, which specifically inhibits mTOR. Furthermore, we observed that rapamycin displays a potent antitumor effect in vivo, as it inhibits DNA synthesis and induces the apoptotic death of HNSCC cells, ultimately resulting in tumor regression. These findings identify the Akt-mTOR pathway as a potential therapeutic target for HNSCC, and may provide the rationale for the early clinical evaluation of rapamycin and its analogues in patients with HNSCC.
High Frequencies of Functional Tumor-Reactive T Cells in Bone Marrow and Blood of Pancreatic Cancer Patients
Authors: Schmitz-Winnenthal Friedrich H. , Volk Christine , Z'graggen Kaspar , Galindo Luis , Nummer Daniel , Ziouta Yvonne , Bucur Marianna , Weitz Jürgen , Schirrmacher Volker , Büchler Markus W. , Beckhove Philipp 
Published: November 2005

Abstract
Pancreatic cancer is characterized by aggressive growth and treatment resistance. New approaches include immunotherapeutic strategies but the type and extent of spontaneous immune responses against tumor antigens remains unclear. A dominance of TH2 cytokines in patients' sera reported previously suggests systemic tumor-induced immunosuppression, potentially inhibiting the induction of tumor-reactive T cells. We characterized the localization, frequencies, and functional potential of spontaneously induced memory T cells specific for individual tumor antigens or the tumor-associated antigen mucin-1 in the peripheral blood and bone marrow of 41 pancreatic cancer patients. We found high numbers of tumor-reactive T cells in all bone marrow samples and in 50% of the blood samples. These cells secreted the TH1 cytokine IFN-γ rather than TH2 cytokines upon stimulation with tumor antigens. Although consistently induced during pancreatic cancer, T cells specific for pancreatic antigens were not detected during chronic pancreatitis, suggesting that their evaluation may be of diagnostic use in both diseases. Freshly isolated T cells from cancer patients recognized autologous tumor cells and rejected them in vitro and in a xenotransplant model in vivo, suggesting their therapeutic potential. Thus, tumor antigen–specific T cell responses occur regularly during pancreatic cancer disease and lead to enrichment of tumor cell–reactive memory T cells in the bone marrow. The bone marrow can therefore be considered an important organ for antitumor immune responses in pancreatic cancer.
Loss of Heterozygosity and Somatic Mutations of the Glucocorticoid Receptor Gene Are Rarely Found at Relapse in Pediatric Acute Lymphoblastic Leukemia but May Occur in a Subpopulation Early in the Disease Course
Authors: Irving Julie A.E. , Minto Lynne , Bailey Simon , Hall Andy G. 
Published: November 2005

Abstract
Glucocorticoids are pivotal in the treatment of children with acute lymphoblastic leukemia (ALL) and have significant antileukemic effects in the majority of children. However, clinical resistance is a significant problem. Although cell line models implicate somatic mutations and loss of heterozygosity (LOH) of the glucocorticoid receptor (GR) gene as a mechanism of in vitro glucocorticoid resistance, the relevance of this mechanism as a cause of clinical resistance in children with ALL is not known. Mutational screening of all coding exons of the GR gene and LOH analyses were done in a large cohort of relapsed ALL. We show that somatic mutations and LOH of the GR rarely contribute to relapsed disease in children with ALL. However, we report the second case of ALL with a somatic mutation of the GR involving a 29-bp deletion in exon 8 and resulting in a truncated protein with loss of part of the ligand-binding domain. There was no evidence of a remaining wild-type allele. Allele-specific PCR detected the mutated clone at day 28 after presentation, which persisted at a low level throughout the disease course before relapse several years later. We hypothesize that the mutated allele present in a leukemic subclone at initial diagnosis was selected for during remission induction with glucocorticoids and contributed to the emergence of a glucocorticoid-resistant cell population.
Application of Embryonic Lethal or Other Obvious Phenotypes to Characterize the Clinical Significance of Genetic Variants Found in Trans with Known Deleterious Mutations
Authors: Judkins Thaddeus , Hendrickson Brant C. , Deffenbaugh Amie M. , Eliason Kristilyn , Leclair Benoît , Norton Michael J. , Ward Brian E. , Pruss Dmitri , Scholl Thomas 
Published: November 2005

Abstract
This work describes an approach to characterize the clinical significance of genetic variants detected during the genetic testing of BRCA1 in patients from hereditary breast/ovarian cancer families. Results from transgenic mice and extensive clinical testing support the hypothesis that biallelic BRCA1 mutations result in embryonic lethality. Therefore, it is reasonable to conclude that variants of uncertain clinical significance found to reside in trans with known deleterious mutations impart reduced risk for cancer. This approach was applied to a large data set of 55,630 patients who underwent clinical BRCA1 screening by whole gene direct DNA sequencing. Fourteen common single nucleotide polymorphisms (SNPs) were used to assign 10 previously defined common, recurrent, or canonical haplotypes in 99% of these cases. From a total of 1,477 genetic variants detected in these patients, excluding haplotype-tagging SNPs, 877 (59%) could be unambiguously assigned to one or more haplotypes. In 41 instances, variants previously classified as being of uncertain clinical significance, mostly missense variants, were excluded as fully penetrant mutations due to their coincidence in trans with known deleterious mutations. From a total of 1,150 patients that harbored these 41 variants, 956 carried one as the sole variant of uncertain clinical significance reported. This approach could have widespread application to other disease genes where compound heterozygous mutations are incompatible with life or result in obvious phenotypes. This largely computational technique is advantageous because it relies upon existing clinical data and is likely to prove informative for prevalent genetic variants in large data sets.
Retinol Inhibits the Growth of All-Trans-Retinoic Acid–Sensitive and All-Trans-Retinoic Acid–Resistant Colon Cancer Cells through a Retinoic Acid Receptor–Independent Mechanism
Authors: Park Eun Young , Dillard Alice , Williams Elizabeth A. , Wilder Erik T. , Pepper M. Reese , Lane Michelle A. 
Published: November 2005

Abstract
Retinol (vitamin A) is thought to exert its effects through the actions of its metabolite, all-trans-retinoic acid (ATRA), on gene transcription mediated by retinoic acid receptors (RAR) and retinoic acid response elements (RARE). However, retinoic acid resistance limits the chemotherapeutic potential of ATRA. We examined the ability of retinol to inhibit the growth of ATRA-sensitive (HCT-15) and ATRA-resistant (HCT-116, SW620, and WiDR) human colon cancer cell lines. Retinol inhibited cell growth in a dose-responsive manner. Retinol was not metabolized to ATRA or any bioactive retinoid in two of the cell lines examined. HCT-116 and WiDR cells converted a small amount of retinol to ATRA; however, this amount of ATRA was unable to inhibit cell growth. To show that retinol was not inducing RARE-mediated transcription, each cell line was transfected with pRARE-chloramphenicol acetyltransferase (CAT) and treated with ATRA and retinol. Although treatment with ATRA increased CAT activity 5-fold in ATRA-sensitive cells, retinol treatment did not increase CAT activity in any cell line examined. To show that growth inhibition due to retinol was ATRA, RAR, and RARE independent, a pan-RAR antagonist was used to block RAR signaling. Retinol-induced growth inhibition was not alleviated by the RAR antagonist in any cell line, but the antagonist alleviated ATRA-induced growth inhibition of HCT-15 cells. Retinol did not induce apoptosis, differentiation or necrosis, but affected cell cycle progression. Our data show that retinol acts through a novel, RAR-independent mechanism to inhibit colon cancer cell growth.
Epigenetic Regulation of WTH3 in Primary and Cultured Drug-Resistant Breast Cancer Cells
Authors: Tian Kegui , Jurukovski Vladimir , Wang Xue-Ping , Kaplan Mark H. , Xu Haopeng 
Published: November 2005

Abstract
Previous studies showed that the WTH3 gene functioned as a negative regulator during multidrug resistance (MDR) development in vitro. To understand whether this gene is also involved in clinical drug resistance, hypermethylation at its promoter region observed in cultured MDR MCF7/AdrR cells was examined in primary drug-resistant breast cancer epithelial cells isolated from effusions of breast cancer patients. The results showed that this event also occurred in drug-resistant breast cancer epithelial cells and a newly induced drug-resistant cell line, MCF7/inR. Interestingly, we found that a CpG (CpG 23) that was close to the TATA-like box was constantly methylated in the WTH3 promoter of drug-resistant breast cancer epithelial and cultured MDR cells. Mutagenic study suggested that this CpG site had a functional effect on promoter activity. We also discovered that MCF7/AdrR cells treated with trichostatin A, a histone deacetylase inhibitor, exhibited higher WTH3, but lower MDR1, expression. A reverse correlation between WTH3 and MDR1 gene expression was also observed in MCF7/AdrR, and its non-MDR parental cell line, MCF7/WT. This result indicated that both DNA methylation and histone deacetylase could act in concert to inhibit WTH3 and consequently stimulate MDR1 expression. This hypothesis was supported by data obtained from introducing the WTH3 transgene into MDR cell lines, which reduced endogenous MDR1 expression. Therefore, our studies suggested that the behavior of WTH3 in primary drug-resistant breast cancer epithelial cells was similar to that in a model system where epigenetic regulation of the WTH3 gene was linked to the MDR phenotype.