bluuebunny/crossref_metadata_2025_split huggingface dataset:
Counting Legionella Cells within Single Amoeba Host Cells
Authors: Buse Helen Y. , Ashbolt Nicholas J. 
Published: January 2012

ABSTRACT

Here we present the first attempt to quantify
Legionella pneumophila
cell numbers within individual amoeba hosts that may be released into engineered water systems. The maximum numbers of culturable
L. pneumophila
cells grown within
Acanthamoeba polyphaga
and
Naegleria fowleri
were 1,348 (mean, 329) and 385 (mean, 44) CFU trophozoite
−1
, respectively.
Regulation of Bacteriocin Production and Cell Death by the VicRK Signaling System in Streptococcus mutans
Authors: Senadheera D. B. , Cordova M. , Ayala E. A. , Chávez de Paz L. E. , Singh K. , Downey J. S. , Svensäter G. , Goodman S. D. , Cvitkovitch D. G. 
Published: January 2012

ABSTRACT

The VicRK two-component signaling system modulates biofilm formation, genetic competence, and stress tolerance in
Streptococcus mutans
. We show here that the VicRK modulates bacteriocin production and cell viability, in part by direct modulation of competence-stimulating peptide (CSP) production in
S. mutans
. Global transcriptome and real-time transcriptional analysis of the VicK-deficient mutant (SmuvicK) revealed significant modulation of several bacteriocin-related loci, including
nlmAB
,
nlmC
, and
nlmD
(
P
< 0.001), suggesting a role for the VicRK in producing mutacins IV, V, and VI. Bacteriocin overlay assays revealed an altered ability of the
vic
mutants to kill related species. Since a well-conserved VicR binding site (TGTWAH-N
5
-TGTWAH) was identified within the
comC
coding region, we confirmed VicR binding to this sequence using DNA footprinting. Overexpression of the
vic
operon caused growth-phase-dependent repression of
comC
,
comDE
, and
comX.
In the
vic
mutants, transcription of
nlmC/cipB
encoding mutacin V, previously linked to CSP-dependent cell lysis, as well as expression of its putative immunity factor encoded by
immB
, were significantly affected relative to the wild type (
P
< 0.05). In contrast to previous reports that proposed a hyper-resistant phenotype for the VicK mutant in cell viability, the release of extracellular genomic DNA was significantly enhanced in SmuvicK (
P
< 0.05), likely as a result of increased autolysis compared with the parent. The drastic influence of VicRK on cell viability was also demonstrated using
vic
mutant biofilms. Taken together, we have identified a novel regulatory link between the VicRK and ComDE systems to modulate bacteriocin production and cell viability of
S. mutans
.
Characterization and Functional Analysis of
atl
, a Novel Gene Encoding Autolysin in Streptococcus suis
Authors: Ju Cun-Xiang , Gu Hong-Wei , Lu Cheng-Ping 
Published: January 2012

ABSTRACT

Streptococcus suis
serotype 2 (
S. suis
2) is an important swine and human pathogen responsible for septicemia and meningitis. A novel gene, designated
atl
and encoding a major autolysin of
S. suis
2 virulent strain HA9801, was identified and characterized in this study. The Atl protein contains 1,025 amino acids with a predicted molecular mass of 113 kDa and has a conserved
N
-acetylmuramoyl-
l
-alanine amidase domain. Recombinant Atl was expressed in
Escherichia coli
, and its bacteriolytic and fibronectin-binding activities were confirmed by zymography and Western affinity blotting. Two bacteriolytic bands were shown in the sodium dodecyl sulfate extracts of HA9801, while both were absent from the
atl
inactivated mutant. Cell chains of the mutant strain became longer than that of the parental strain. In the autolysis assay, HA9801 decreased to 20% of the initial optical density (OD) value, while the mutant strain had almost no autolytic activity. The biofilm capacity of the
atl
mutant was reduced ∼30% compared to the parental strain. In the zebrafish infection model, the 50% lethal dose of the mutant strain was increased up to 5-fold. Furthermore, the adherence to HEp-2 cells of the
atl
mutant was 50% less than that of the parental strain. Based on the functional analysis of the recombinant Atl and observed effects of
atl
inactivation on HA9801, we conclude that Atl is a major autolysin of HA9801. It takes part in cell autolysis, separation of daughter cells, biofilm formation, fibronectin-binding activity, cell adhesion, and pathogenesis of HA9801.
Legionella pneumophila LidA Affects Nucleotide Binding and Activity of the Host GTPase Rab1
Authors: Neunuebel M. Ramona , Mohammadi Sina , Jarnik Michal , Machner Matthias P. 
Published: January 2012

ABSTRACT

Legionella pneumophila
, the causative agent of a severe pneumonia known as Legionnaires' disease, intercepts material from host cell membrane transport pathways to create a specialized vacuolar compartment that supports bacterial replication. Delivery of bacterial effector proteins into the host cell requires the Dot/Icm type IV secretion system. Several effectors, including SidM, SidD, and LepB, were shown to target the early secretory pathway by manipulating the activity of the host GTPase Rab1. While the function of these effectors has been well characterized, the role of another Rab1-interacting protein from
L. pneumophila
, the effector protein LidA, is poorly understood. Here, we show that LidA binding to Rab1 stabilized the Rab1-guanosine nucleotide complex, protecting it from inactivation by GTPase-activating proteins (GAPs) and from nucleotide extraction. The protective effect of LidA on the Rab1-guanine nucleotide complex was concentration dependent, consistent with a 1:1 stoichiometry of the LidA-Rab1 complex. The central coiled-coil region of LidA was sufficient for Rab1 binding and to prevent GAP-mediated inactivation or nucleotide extraction from Rab1. In addition, the central region mediated binding to phosphatidylinositol 3-phosphate and other phosphoinositides. When bound to Rab1, LidA interfered with the covalent modification of Rab1 by phosphocholination or AMPylation, and it also blocked de-AMPylation of Rab1 by SidD and dephosphocholination by Lem3. Based on these findings, we propose a role for LidA in bridging the membrane of the
Legionella
-containing vacuole (LCV) with that of secretory transport vesicles surrounding the LCV.
Physiology of Resistant Deinococcus geothermalis Bacterium Aerobically Cultivated in Low-Manganese Medium
Authors: Liedert Christina , Peltola Minna , Bernhardt Jörg , Neubauer Peter , Salkinoja-Salonen Mirja 
Published: January 2012

ABSTRACT

This dynamic proteome study describes the physiology of growth and survival of
Deinococcus geothermalis
, in conditions simulating paper machine waters being aerobic, warm, and low in carbon and manganese. The industrial environment of this species differs from its natural habitats, geothermal springs and deep ocean subsurfaces, by being highly exposed to oxygen. Quantitative proteome analysis using two-dimensional gel electrophoresis and bioinformatic tools showed expression change for 165 proteins, from which 47 were assigned to a function. We propose that
D. geothermalis
grew and survived in aerobic conditions by channeling central carbon metabolism to pathways where mainly NADPH rather than NADH was retrieved from the carbon source. A major part of the carbon substrate was converted into succinate, which was not a fermentation product but likely served combating reactive oxygen species (ROS). Transition from growth to nongrowth resulted in downregulation of the oxidative phosphorylation observed as reduced expression of V-type ATPase responsible for ATP synthesis in
D. geothermalis
. The battle against oxidative stress was seen as upregulation of superoxide dismutase (Mn dependent) and catalase, as well as several protein repair enzymes, including FeS cluster assembly proteins of the iron-sulfur cluster assembly protein system, peptidylprolyl isomerase, and chaperones. Addition of soluble Mn reinitiated respiration and proliferation with concomitant acidification, indicating that aerobic metabolism was restricted by access to manganese. We conclude that
D. geothermalis
prefers to combat ROS using manganese-dependent enzymes, but when manganese is not available central carbon metabolism is used to produce ROS neutralizing metabolites at the expense of high utilization of carbon substrate.
Immunochemotherapy with Intensive Consolidation for Primary CNS Lymphoma: A Pilot Study and Prognostic Assessment by Diffusion-Weighted MRI
Authors: Wieduwilt Matthew J. , Valles Francisco , Issa Samar , Behler Caroline M. , Hwang James , McDermott Michael , Treseler Patrick , O'Brien Joan , Shuman Marc A. , Cha Soonmee , Damon Lloyd E. , Rubenstein James L. 
Published: January 2012

Abstract
Purpose: We evaluated a novel therapy for primary central nervous system lymphoma (PCNSL) with induction immunochemotherapy with high-dose methotrexate, temozolomide, and rituximab (MT-R) followed by intensive consolidation with infusional etoposide and high-dose cytarabine (EA). In addition, we evaluated the prognostic value of the minimum apparent diffusion coefficient (ADCmin) derived from diffusion-weighted MRI (DW-MRI) in patients treated with this regimen.
Experimental Design: Thirty-one patients (median age, 61 years; median Karnofsky performance score, 60) received induction with methotrexate every 14 days for 8 planned cycles. Rituximab was administered the first 6 cycles and temozolomide administered on odd-numbered cycles. Patients with responsive or stable central nervous system (CNS) disease received EA consolidation. Pretreatment DW-MRI was used to calculate the ADCmin of contrast-enhancing lesions.
Results: The complete response rate for MT-R induction was 52%. At a median follow-up of 79 months, the 2-year progression-free and overall survival were 45% and 58%, respectively. For patients receiving EA consolidation, the 2-year progression-free and overall survival were 78% and 93%, respectively. EA consolidation was also effective in an additional 3 patients who presented with synchronous CNS and systemic lymphoma. Tumor ADCmin less than 384 × 10–6 mm2/s was significantly associated with shorter progression-free and overall survival.
Conclusions: MT-R induction was effective and well tolerated. MT-R followed by EA consolidation yielded progression-free and overall survival outcomes comparable to regimens with chemotherapy followed by whole-brain radiotherapy consolidation but without evidence of neurotoxicity. Tumor ADCmin derived from DW-MRI provided better prognostic information for PCNSL patients treated with the MTR-EA regimen than established clinical risk scores. Clin Cancer Res; 18(4); 1146–55. ©2012 AACR.
Notch Signaling Promotes Growth and Invasion in Uveal Melanoma
Authors: Asnaghi Laura , Ebrahimi Katayoon B. , Schreck Karisa C. , Bar Eli E. , Coonfield Michael L. , Bell W. Robert , Handa James , Merbs Shannath L. , Harbour J. William , Eberhart Charles G. 
Published: January 2012

Abstract
Purpose: To determine whether uveal melanoma, the most common primary intraocular malignancy in adults, requires Notch activity for growth and metastasis.
Experimental Design: Expression of Notch pathway members was characterized in primary tumor samples and in cell lines, along with the effects of Notch inhibition or activation on tumor growth and invasion.
Results: Notch receptors, ligands, and targets were expressed in all five cell lines examined and in 30 primary uveal melanoma samples. Interestingly, the three lines with high levels of baseline pathway activity (OCM1, OCM3, and OCM8) had their growth reduced by pharmacologic Notch blockade using the γ-secretase inhibitor (GSI) MRK003. In contrast, two uveal melanoma lines (Mel285 and Mel290) with very low expression of Notch targets were insensitive to the GSI. Constitutively active forms of Notch1 and Notch2 promoted growth of uveal melanoma cultures and were able to rescue the inhibitory effects of GSI. MRK003 treatment also inhibited anchorage-independent clonogenic growth and cell invasion and reduced phosphorylation levels of STAT3 and extracellular signal-regulated kinase (Erk)1/2. Suppression of canonical Notch activity using short hairpin RNA targeting Notch2 or CBF1 was also able to reduce tumor growth and invasion. Finally, intraocular xenograft growth was significantly decreased by GSI treatment.
Conclusion: Our findings suggest that Notch plays an important role in inducing proliferation and invasion in uveal melanoma and that inhibiting this pathway may be effective in preventing tumor growth and metastasis. Clin Cancer Res; 18(3); 654–65. ©2012 AACR.
Regulation of CD20 in Rituximab-Resistant Cell Lines and B-cell Non-Hodgkin Lymphoma
Authors: Tsai Ping-Chiao , Hernandez-Ilizaliturri Francisco J. , Bangia Naveen , Olejniczak Scott H. , Czuczman Myron S. 
Published: January 2012

AbstractPurpose: The aim of this research was to further investigate the contribution of CD20 antigen expression to rituximab activity and define the mechanisms responsible for CD20 downregulation in rituximab-resistant cell lines (RRCL).Experimental Design: Rituximab-sensitive cell lines, RRCL, and primary neoplastic B cells were evaluated by chromium-51 release assays, ImageStream image analysis, immunohistochemical staining, flow cytometric analysis, CD20 knockdown, promoter activity, chromatin immunoprecipitation (ChIP) analysis of CD20 promoter, and CD20 plasmid transfection experiments to identify mechanisms associated with CD20 regulation in RRCL.Results: RRCL exhibited a gradual loss of CD20 surface expression with repeated exposure to rituximab. We identified a CD20 antigen surface threshold level required for effective rituximab-associated complement-mediated cytotoxicity (CMC). However, a direct correlation between CD20 surface expression and rituximab-CMC was observed only in rituximab-sensitive cell lines. CD20 promoter activity was decreased in RRCL. Detailed analysis of various CD20 promoter fragments suggested a lack of positive regulatory factors in RRCL. ChIP analysis showed reduced binding of several key positive regulatory proteins on CD20 promoter in RRCL. Interleukin-4 (IL-4) induced higher CD20 promoter activity and CD20 expression but modestly improved rituximab activity in RRCL and in primary B-cell lymphoma cells. Forced CD20 expression restored cytoplasmic but not surface CD20, suggesting the existence of a defect in CD20 protein transport in RRCL.Conclusions: We identified several mechanisms that alter CD20 expression in RRCL and showed that, whereas CD20 expression is important for rituximab activity, additional factors likely contribute to rituximab sensitivity in B-cell lymphoma. Clin Cancer Res; 18(4); 1039–50. ©2012 AACR.
Phase II Trial of Temozolomide in Patients with Relapsed Sensitive or Refractory Small Cell Lung Cancer, with Assessment of Methylguanine-DNA Methyltransferase as a Potential Biomarker
Authors: Pietanza M. Catherine , Kadota Kyuichi , Huberman Kety , Sima Camelia S. , Fiore John J. , Sumner Dyana K. , Travis William D. , Heguy Adriana , Ginsberg Michelle S. , Holodny Andrei I. , Chan Timothy A. , Rizvi Naiyer A. , Azzoli Christopher G. , Riely Gregory J. , Kris Mark G. , Krug Lee M. 
Published: January 2012

Abstract
Purpose: This phase II study was conducted to assess the efficacy of temozolomide in patients with relapsed small cell lung cancer (SCLC).
Experimental Design: Patients with disease progression after one or two prior chemotherapy regimens received temozolomide at 75 mg/m2/d for 21 days of a 28-day cycle. The primary endpoint was the overall response rate [ORR; complete response (CR) plus partial response (PR)], which was evaluated separately in sensitive and refractory cohorts. In the available tissue, we assessed O6-methylguanine-DNA methyltransferase (MGMT) promoter methylation status by PCR and MGMT expression by immunohistochemistry.
Results: Sixty-four patients were accrued: 48 patients in the sensitive cohort and 16 in the refractory group. One CR and 10 PRs were noted in sensitive patients [ORR, 23%; 95% confidence interval (CI), 12%–37%]. Two PRs were seen in the refractory cohort (ORR, 13%; 95% CI, 2%–38%). As second- and third-line treatment, the ORR was 22% (95% CI, 9%–40%) and 19% (95% CI, 7%–36%), respectively. Among patients with target brain lesions, 38% had a CR or PR (95% CI, 14%–68%). Grade ≥3 thrombocytopenia and neutropenia were observed in nine patients (14%). A greater number of cases with methylated MGMT had a response compared to those with unmethylated MGMT (38% vs. 7%; P = 0.08).
Conclusion: Temozolomide has activity in relapsed SCLC, particularly for brain metastases. Response to temozolomide may correlate with MGMT methylation in SCLC. Clin Cancer Res; 18(4); 1138–45. ©2012 AACR.
Changes in Keratin Expression during Metastatic Progression of Breast Cancer: Impact on the Detection of Circulating Tumor Cells
Authors: Joosse Simon A. , Hannemann Juliane , Spötter Julia , Bauche Andreas , Andreas Antje , Müller Volkmar , Pantel Klaus 
Published: January 2012

AbstractPurpose: Circulating tumor cells (CTC) might function as early markers for breast cancer metastasis or monitoring therapy efficacy. Enrichment and identification of CTCs are based on epithelial markers that might be modulated during epithelial–mesenchymal transition. Little is known about the expression of keratins in CTCs and whether all CTCs can be detected with antibodies directed against a limited panel of keratins.Experimental Design: Protein expression of keratin 2, 4–10, 13–16, 18, and 19 were assessed by a cocktail of antibodies (C11, AE1, AE3, and K7) and keratin antibodies C11 and A45-B/B3 alone in 11 breast cancer cell lines and 50 primary breast carcinomas and their lymph node metastases. Furthermore, CTCs were assessed in blood of 70 metastatic breast cancer patients.Results: Claudin-low cell lines did not show expression of normal breast epithelial keratins but were positive for K14 and K16, detected by the cocktail only. Primary breast carcinomas showed changes in keratin expression during metastatic progression to the lymph nodes. In 35 of 70 patients CTCs were identified, of which 83%, 40%, and 57% were identified by the cocktail, C11 and A45-B/B3, respectively. Identification of CTCs by the cocktail was associated with shorter survival (P &lt; 0.01). In silico analyses revealed association between KRT16 expression and shorter relapse-free survival in metastatic breast cancer.Conclusion: Breast cancer cells show a complex pattern of keratin expression with potential biologic relevance. Individual keratin antibodies recognizing only a limited set of keratins inherit the risk to miss biologically relevant CTCs in cancer patients, and antibody cocktails including these keratins are therefore recommended. Clin Cancer Res; 18(4); 993–1003. ©2012 AACR.
Combination of a Novel Gene Expression Signature with a Clinical Nomogram Improves the Prediction of Survival in High-Risk Bladder Cancer
Authors: Riester Markus , Taylor Jennifer M. , Feifer Andrew , Koppie Theresa , Rosenberg Jonathan E. , Downey Robert J. , Bochner Bernard H. , Michor Franziska 
Published: January 2012

Abstract
Purpose: We aimed to validate and improve prognostic signatures for high-risk urothelial carcinoma of the bladder.
Experimental Design: We evaluated microarray data from 93 patients with bladder cancer managed by radical cystectomy to determine gene expression patterns associated with clinical and prognostic variables. We compared our results with published bladder cancer microarray data sets comprising 578 additional patients and with 49 published gene signatures from multiple cancer types. Hierarchical clustering was utilized to identify subtypes associated with differences in survival. We then investigated whether the addition of survival-associated gene expression information to a validated postcystectomy nomogram utilizing clinical and pathologic variables improves prediction of recurrence.
Results: Multiple markers for muscle invasive disease with highly significant expression differences in multiple data sets were identified, such as fibronectin 1 (FN1), NNMT, POSTN, and SMAD6. We identified signatures associated with pathologic stage and the likelihood of developing metastasis and death from bladder cancer, as well as with two distinct clustering subtypes of bladder cancer. Our novel signature correlated with overall survival in multiple independent data sets, significantly improving the prediction concordance of standard staging in all data sets [mean ΔC-statistic: 0.14; 95% confidence interval (CI), 0.01–0.27; P &lt; 0.001]. Tested in our patient cohort, it significantly enhanced the performance of a postoperative survival nomogram (ΔC-statistic: 0.08, 95% CI, −0.04–0.20; P &lt; 0.005).
Conclusions: Prognostic information obtained from gene expression data can aid in posttreatment prediction of bladder cancer recurrence. Our findings require further validation in external cohorts and prospectively in a clinical trial setting. Clin Cancer Res; 18(5); 1323–33. ©2012 AACR.
Detection of TIMP3 Promoter Hypermethylation in Salivary Rinse as an Independent Predictor of Local Recurrence-Free Survival in Head and Neck Cancer
Authors: Sun Wenyue , Zaboli David , Wang Hao , Liu Yan , Arnaoutakis Demetri , Khan Tanbir , Khan Zubair , Koch Wayne M. , Califano Joseph A. 
Published: January 2012

Abstract
Purpose: To validate a panel of methylation-based salivary rinse biomarkers (P16, CCNA1, DCC, TIMP3, MGMT, DAPK, and MINT31) previously shown to be independently associated with poor overall survival and local recurrence in a larger, separate cohort of patients with head and neck squamous cell carcinoma (HNSCC).
Experimental Design: One hundred ninety-seven patients were included. All pretreatment saliva DNA samples were evaluated for the methylation status of the gene promoters by quantitative methylation-specific PCR. The main outcome measures were overall survival, local recurrence-free survival, and disease-free survival.
Results: In univariate analyses, the detection of hypermethylation of CCNA1, MGMT, and MINT31 was significantly associated with poor overall survival; the detection of hypermethylation of TIMP3 was significantly associated with local recurrence-free survival; and the detection of hypermethylation of MINT31 was significantly associated with poor disease-free survival. In multivariate analyses, detection of hypermethylation at any single marker was not predictive of overall survival in patients with HNSCC; detection of hypermethylation of TIMP3 in salivary rinse had an independent, significant association with local recurrence-free survival (HR = 2.51; 95% CI: 1.10–5.68); and none of the studied markers was significantly associated with disease-free survival.
Conclusion: The detection of promoter hypermethylation of the seven genes in salivary rinse as an independent prognostic indicator of overall survival in patients with HNSCC was not validated. Detection of promoter hypermethylation of TIMP3 in pretreatment salivary rinse is independently associated with local recurrence-free survival in patients with HNSCC and may be a valuable salivary rinse biomarker for HNSCC recurrence. Clin Cancer Res; 18(4); 1082–91. ©2012 AACR.