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Huang L.,University of Florida | Gregory J.F.,III
Nature Chemical Biology | Year: 2016

DUF89 family proteins occur widely in both prokaryotes and eukaryotes, but their functions are unknown. Here we define three DUF89 subfamilies (I, II, and III), with subfamily II being split into stand-alone proteins and proteins fused to pantothenate kinase (PanK). We demonstrated that DUF89 proteins have metal-dependent phosphatase activity against reactive phosphoesters or their damaged forms, notably sugar phosphates (subfamilies II and III), phosphopantetheine and its S-sulfonate or sulfonate (subfamily II-PanK fusions), and nucleotides (subfamily I). Genetic and comparative genomic data strongly associated DUF89 genes with phosphoester metabolism. The crystal structure of the yeast (Saccharomyces cerevisiae) subfamily III protein YMR027W revealed a novel phosphatase active site with fructose 6-phosphate and Mg2+ bound near conserved signature residues Asp254 and Asn255 that are critical for activity. These findings indicate that DUF89 proteins are previously unrecognized hydrolases whose characteristic in vivo function is to limit potentially harmful buildups of normal or damaged phosphometabolites. © 2016 Nature Publishing Group, a division of Macmillan Publishers Limited. All Rights Reserved.

Merker M.,Research Center Borstel | Barry C.E.,III
Nature Genetics | Year: 2015

Mycobacterium tuberculosis strains of the Beijing lineage are globally distributed and are associated with the massive spread of multidrug-resistant (MDR) tuberculosis in Eurasia. Here we reconstructed the biogeographical structure and evolutionary history of this lineage by genetic analysis of 4,987 isolates from 99 countries and whole-genome sequencing of 110 representative isolates. We show that this lineage initially originated in the Far East, from where it radiated worldwide in several waves. We detected successive increases in population size for this pathogen over the last 200 years, practically coinciding with the Industrial Revolution, the First World War and HIV epidemics. Two MDR clones of this lineage started to spread throughout central Asia and Russia concomitantly with the collapse of the public health system in the former Soviet Union. Mutations identified in genes putatively under positive selection and associated with virulence might have favored the expansion of the most successful branches of the lineage. © 2015 Nature Publishing Group, a division of Macmillan Publishers Limited. All Rights Reserved.

Shenai S.,Center for Emerging Pathogens | Barry C.E.,III
Nature Medicine | Year: 2016

The absence of a gold standard to determine when antibiotics induce a sterilizing cure has confounded the development of new approaches to treat pulmonary tuberculosis (PTB). We detected positron emission tomography and computerized tomography (PET–CT) imaging response patterns consistent with active disease, along with the presence of Mycobacterium tuberculosis (MTB) mRNA in sputum and bronchoalveolar lavage samples, in a substantial proportion of adult, HIV-negative patients with PTB after a standard 6-month treatment plus 1 year follow-up, including patients with a durable cure and others who later developed recurrent disease. The presence of MTB mRNA in the context of nonresolving and intensifying lesions on PET–CT images might indicate ongoing transcription, suggesting that even apparently curative treatment for PTB may not eradicate all of the MTB bacteria in most patients. This suggests an important complementary role for the immune response in maintaining a disease-free state. Sterilizing drugs or host-directed therapies, and better treatment response markers, are probably needed for the successful development of improved and shortened PTB-treatment strategies. © 2016 Nature Publishing Group, a division of Macmillan Publishers Limited. All Rights Reserved.

Ahlstrom L.S.,University of Michigan | Brooks C.L.,III
Nature Structural and Molecular Biology | Year: 2016

Challenges in determining the structures of heterogeneous and dynamic protein complexes have greatly hampered past efforts to obtain a mechanistic understanding of many important biological processes. One such process is chaperone-assisted protein folding. Obtaining structural ensembles of chaperone–substrate complexes would ultimately reveal how chaperones help proteins fold into their native state. To address this problem, we devised a new structural biology approach based on X-ray crystallography, termed residual electron and anomalous density (READ). READ enabled us to visualize even sparsely populated conformations of the substrate protein immunity protein 7 (Im7) in complex with the Escherichia coli chaperone Spy, and to capture a series of snapshots depicting the various folding states of Im7 bound to Spy. The ensemble shows that Spy-associated Im7 samples conformations ranging from unfolded to partially folded to native-like states and reveals how a substrate can explore its folding landscape while being bound to a chaperone. © 2016 Nature Publishing Group, a division of Macmillan Publishers Limited. All Rights Reserved.

Poruk K.E.,Sloan Kettering Cancer Center | Valero V.,III
Annals of Surgery | Year: 2016

OBJECTIVE:: We assessed circulating tumor cells (CTCs) with epithelial and mesenchymal phenotypes as a potential prognostic biomarker for patients with pancreatic adenocarcinoma (PDAC). BACKGROUND:: PDAC is the fourth leading cause of cancer death in the United States. There is an urgent need to develop biomarkers that predict patient prognosis and allow for better treatment stratification. METHODS:: Peripheral and portal blood samples were obtained from 50 patients with PDAC before surgical resection and filtered using the Isolation by Size of Epithelial Tumor cells method. CTCs were identified by immunofluorescence using commercially available antibodies to cytokeratin, vimentin, and CD45. RESULTS:: Thirty-nine patients (78%) had epithelial CTCs that expressed cytokeratin but not CD45. Twenty-six (67%) of the 39 patients had CTCs which also expressed vimentin, a mesenchymal marker. No patients had cytokeratin-negative and vimentin-positive CTCs. The presence of cytokeratin-positive CTCs (P< 0.01), but not mesenchymal-like CTCs (P= 0.39), was associated with poorer survival. The presence of cytokeratin-positive CTCs remained a significant independent predictor of survival by multivariable analysis after accounting for other prognostic factors (P< 0.01). The detection of CTCs expressing both vimentin and cytokeratin was predictive of recurrence (P= 0.01). Among patients with cancer recurrence, those with vimentin-positive and cytokeratin-expressing CTCs had decreased median time to recurrence compared with patients without CTCs (P= 0.02). CONCLUSIONS:: CTCs are an exciting potential strategy for understanding the biology of metastases, and provide prognostic utility for PDAC patients. CTCs exist as heterogeneous populations, and assessment should include phenotypic identification tailored to characterize cells based on epithelial and mesenchymal markers. Copyright © 2016 Wolters Kluwer Health, Inc. All rights reserved.

Marakalala M.J.,Harvard University | Barry C.E.,III
Nature Medicine | Year: 2016

Granulomas are the pathological hallmark of tuberculosis (TB). However, their function and mechanisms of formation remain poorly understood. To understand the role of granulomas in TB, we analyzed the proteomes of granulomas from subjects with tuberculosis in an unbiased manner. Using laser-capture microdissection, mass spectrometry and confocal microscopy, we generated detailed molecular maps of human granulomas. We found that the centers of granulomas have a pro-inflammatory environment that is characterized by the presence of antimicrobial peptides, reactive oxygen species and pro-inflammatory eicosanoids. Conversely, the tissue surrounding the caseum has a comparatively anti-inflammatory signature. These findings are consistent across a set of six human subjects and in rabbits. Although the balance between systemic pro- and anti-inflammatory signals is crucial to TB disease outcome, here we find that these signals are physically segregated within each granuloma. From the protein and lipid snapshots of human and rabbit lesions analyzed here, we hypothesize that the pathologic response to TB is shaped by the precise anatomical localization of these inflammatory pathways during the development of the granuloma. © 2016 Nature Publishing Group, a division of Macmillan Publishers Limited. All Rights Reserved.

Nussbaum D.P.,Duke University | Blazer D.G.,III
Journal of the American College of Surgeons | Year: 2015

Background The management of 1- to 2-cm appendiceal carcinoid tumors remains controversial. Here we use the National Cancer Data Base (NCDB) to compare long-term outcomes for patients treated via resection of the primary tumor alone vs right hemicolectomy (RHC). Study Design The 1998 to 2011 NCDB User File was queried to identify patients with 1- to 2-cm appendiceal carcinoids. Patients were stratified by surgical technique: Resection of the primary tumor alone vs RHC with regional lymphadenectomy. Multivariable logistic regression was used to compare short-term outcomes. Survival was estimated using the Kaplan-Meier method with comparisons based on the log-rank test. Results A total of 916 patients were identified, including 42% managed with primary resection and 58% with RHC. Patients who underwent RHC had slightly larger tumors and higher-stage tumors; otherwise, there were no baseline differences between groups. The rates of positive margins were similar (5.5% vs 4.5%; p = 0.60). Among all patients, 1- and 5-year survival were 98.1% and 88.7% vs 96.7% and 87.4% (p = 0.52) for those managed via primary resection vs RHC, respectively. Among patients with moderate/high-grade/anaplastic carcinoids, 1- and 5-year survival were 93.3% and 72.0% vs 92.3% and 71.9%, respectively (p = 0.78). After adjustment with Cox proportional hazards modeling, we confirmed that there was no survival benefit for patients undergoing RHC (hazard ratio = 1.14; p = 0.72). Conclusions For 1- to 2-cm appendiceal carcinoids, formal resection of the right colon does not appear to improve survival, even for higher-grade tumors. Our findings suggest that resection of the primary tumor alone is adequate for all carcinoids <2 cm. © 2015 American College of Surgeons.

Quillin R.C.,III | Shah S.A.,University of Cincinnati
Journal of the American College of Surgeons | Year: 2015

Background Although donation after cardiac death (DCD) liver allografts have been used to expand the donor pool, concerns exist regarding primary nonfunction and biliary complications. Our aim was to compare resource use and outcomes of DCD allografts with donation after brain death (DBD) liver allografts. Study Design Using a linkage between the University HealthSystem Consortium and Scientific Registry of Transplant Recipients databases, we identified 11,856 patients who underwent deceased donor liver transplantation (LT) from 2007 to 2011. Patients were divided into 2 cohorts based on type of allograft (DCD vs DBD). Matched pair analysis (n = 613 in each group) was used to compare outcomes of the 2 donor types. Results Donation after cardiac death allografts comprised 5.2% (n = 613) of all LTs in the studied cohort; DCD allograft recipients were healthier and had lower median Model of End-Stage Liver Disease (MELD) score (17 vs 19; p < 0.0001). Post LT, there was no significant difference in length of stay, perioperative mortality, and discharge to home rates. However, DCD allografts were associated with higher direct cost ($110,414 vs $99,543; p < 0.0001) and 30-day readmission rates (46.4% vs 37.1%; p < 0.0001). Matched analysis revealed that DCD allografts were associated with higher direct cost, readmission rates, and inferior graft survival. Conclusions While confirming the previous reports of inferior graft survival associated with DCD allografts, this is the first national report to show increased financial and resource use associated with DCD compared with DBD allografts in a matched recipient cohort. © 2015 American College of Surgeons.

Doswell C.A.,III
Atmospheric Research | Year: 2015

Basic issues associated with how a forecast becomes effective in helping users make decisions based on weather information are described, with a special emphasis on how this might develop in Europe. The notion of a chain of events that begins when the forecast is issued and ends with the user taking effective actions is used to point out what needs to be done to make the process work properly. Geophysical hazard risks and how people respond to the risks associated with them are discussed, concluding that complacency is a major challenge to helping people make appropriate decisions when severe convective storms threaten them. The situation in Europe regarding the threat of severe convective storms is reviewed and some conclusions are drawn. The key conclusion is that there must be a substantial effort to convince Europeans that they are not immune to severe convective weather hazards, since without public support, the weather community in Europe can do little to mitigate the threats posed by severe convective storms. © 2014 Elsevier B.V.

Goode J.C.,III | McCartney J.S.,University of California at San Diego
Journal of Geotechnical and Geoenvironmental Engineering | Year: 2015

This study presents the results from physical modeling experiments on centrifuge-scale energy foundations in dry sand and unsaturated silt layers. These experiments were performed to characterize end restraint effects on soil-structure interaction for energy foundations in different soils and include tests on foundations with semifloating and end-bearing toe boundary conditions and free-expansion and restrained-expansion head boundary conditions. Two scale-model energy foundations having different lengths were constructed from reinforced concrete to simulate end-bearing and semifloating conditions in soil layers having the same thickness. The foundations include embedded thermocouples and strain gauges, which were calibrated under applied mechanical loads and nonisothermal conditions before testing. The variables measured during the experiments include axial strain and temperature distributions in the foundation, temperature, and volumetric water content measurements in the soil, vertical displacements of the foundation head and soil surface, and axial stress at the foundation head. These variables were used to calculate the distributions in thermal axial stress and thermal axial displacement, which are useful in evaluating soil-structure interaction mechanisms. The results confirm observations from full-scale energy foundations in the field for end-bearing foundations and provide new insight into the behavior of semifloating foundations. Heating of the semifloating foundations in compacted silt led to a clear increase in ultimate capacity, potentially due to changes in radial normal stress and thermally induced water flow, while heating of the semifloating foundations in dry sand led to a negligible change in ultimate capacity. © 2015 American Society of Civil Engineers.

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