Developmental phosphoproteomics identifies the kinase CK2 as a driver of Hedgehog signaling and a therapeutic target in medulloblastoma

A major limitation of targeted cancer therapy is the rapid emergence of drug resistance, which often arises through mutations at or downstream of the drug target or through intrinsic resistance of subpopulations of tumor cells. Medulloblastoma (MB), the most common pediatric brain tumor, is no exception, and MBs that are driven by sonic hedgehog (SHH) signaling are particularly aggressive and drug-resistant. To find new drug targets and therapeutics for MB that may be less susceptible to common resistance mechanisms, we used a developmental phosphoproteomics approach in murine granule neuron precursors (GNPs), the developmental cell of origin of MB. The protein kinase CK2 emerged as a driver of hundreds of phosphorylation events during the proliferative, MB-like stage of GNP growth, including the phosphorylation of three of the eight proteins commonly amplified in MB. CK2 was critical to the stabilization and activity of the transcription factor GLI2, a late downstream effector in SHH signaling. CK2 inhibitors decreased the viability of primary SHH-type MB patient cells in culture and blocked the growth of murine MB tumors that were resistant to currently available Hh inhibitors, thereby extending the survival of tumor-bearing mice. Because of structural interactions, one CK2 inhibitor (CX-4945) inhibited both wild-type and mutant CK2, indicating that this drug may avoid at least one common mode of acquired resistance. These findings suggest that CK2 inhibitors may be effective for treating patients with MB and show how phosphoproteomics may be used to gain insight into developmental biology and pathology

Twin Related Domains in Polycrystalline Nickel-Base Superalloys: 3D Structure and Fatigue

Fatigue is the life limiting property of polycrystalline nickel-base superalloys used in turbine disks in the hot section of turbine engines for aerospace and power generation applications. Fabricating components through a powder metallurgy route provides enhanced properties and removes many extrinsic defects associated with fatigue crack formation, driving cracks to initiate at intrinsic microstructrual extremes. Fatigue cracks in the powder metallurgy alloy René 88DT frequently initiate in large grains that are in the tail of the size distribution and contain favorably oriented annealing twin boundaries. Fully characterizing twin boundaries and twin related domains requires 3D microstructural volumes. Datasets of 3D microstructure for René 88DT were collected by TriBeam tomography across a range of resolutions and volumes to characterize microstructure and annealing twins. Algorithms developed to analyze these datasets allow identification of strain localizing sites from 2D cross sections and the frequency of microstructural features amenable to fatigue crack initiation is statistically assessed. Twin related domain structure is quantified via network analysis and leveraged to propose a criterion for identifying fatal fatigue crack sites

Probabilistic Protein Design, Comparative Modeling, and the Structure of a Multidomain P53 Oligomer Bound to DNA

Proteins are the main functional components of all cellular processes, and most of them fold into unique three-dimensional shapes guided by their amino-acid sequence. Discovering the structure of a protein, or protein complexes, can provide important clues about how they perform their function. However, the chemical, physical or architectural properties of many proteins impede traditional approaches to structure determination. Two such proteins, the tumor suppressor p53 and the cholesterol processing enzyme endothelial lipase, are prime examples of problematic proteins that defy structural investigation via crystallographic methods. Therefore, new techniques must be developed to gain valuable structural insights, such as: computationally assisted protein design strategies, more efficient crystal screening, or a combination of both. We applied a statistical computationally assisted design strategy to stabilize a p53 variant consisting of two independently folding domains. The re-engineered variant retained normal DNA-binding activities, and allowed us to experimentally determine the first structure of a physiologically active multi-domain p53 tetramer bound to a full-length DNA response element. We then demonstrated how computational methodology can be used to gain functional detail of proteins in the absence of experimentally determined structures. By creating comparative models of endothelial lipase, we discovered structural features that describe function and regulation, and gained a better understanding of the mechanisms conferring substrate specificity. Additionally, traditional methods for protein structure determination, such as X-ray crystallography, require relatively large amounts of purified sample in order to screen a sufficient variety of conditions. To improve this process, we developed a novel method for protein crystal screening using a microfluidics platform. We show how it is possible to use smaller quantities of protein to screen larger varieties of conditions, in turn increasing the probability of success in obtaining crystals. Furthermore, in contrast to current crystallographic approaches, all steps from screening to crystal growth to data collection were performed within the same reaction chamber, without any manipulation of the crystal, dramatically increasing the efficiency of both time and sample required to realize the structure. Collectively, these results demonstrate how advances in computational and experimental approaches can provide structural detail for proteins in circumstances where traditional methodology fails

2018 GREAT Day Program

SUNY Geneseo’s Twelfth Annual GREAT Day.https://knightscholar.geneseo.edu/program-2007/1012/thumbnail.jp

Integrative modelling of angiogenesis in the bovine corpus luteum

The corpus luteum (CL) is a tissue formed from the remnants of an ovulated follicle in the ovary, and it produces the progesterone needed for a healthy pregnancy. CL growth is highly dependent on a growing nutrient supply, and can be compared with the most aggressive vascular tumours. Angiogenesis, the growth of new blood vessels from existing ones, plays a key role in the growth and function of the CL. Inadequate angiogenesis has been linked to infertility in cows. The CL is composed of several vascular(e.g. endothelial cells (ECs), pericytes (PCs)), and avascular (e.g. luteal cells (LCs), immune cells) cell types, and several pro-angiogenic factors (e.g. Fibroblast Growth Factor 2, FGF2) found to be important in the angiogenic process. The objective of this thesis is to shed light on the cellular and extracellular level determinants of angiogenesis in the bovine CL. We begin with the relevant biological and mathematical literature in Chapter 1. In Chapter 2, an ordinary differential equation model of CL growth is introduced. We assume that the CL volume is a continuum of three cell types, ECs, LCs, and stromal cells (such as PCs). The fourth variable in the model, FGF2, enhances the EC proliferation rate. The model is able, by varying parameters such as the maximal proliferation rate of the ECs, to distinguish cases where the CL shifts from a ‘normal’ to a ‘pathological’ growth. In Chapter 3, we present in vitro CL published and novel studies from Robinson’s Lab. Preliminary results demonstrate interesting endothelial and pericyte behaviours regarding cell aggregation and sprout formation, which are the motivation for the next two Chapters. In these experimental studies, all the CL cell types were incorporated in the same in vitro culture, hence providing a closer approximation to the in vivo environment compared to other in vitro cultures which use only a single cell type (mainly ECs). However, this complicates matters in terms of distinguishing cell behaviours and factors which contribute on the overall cell dynamics. Therefore, in the Chapters 4 and 5 we use data from literature. In Chapter 4, by using the Cellular Potts Model (CPM) framework, we focus on EC-PC interactions, and particularly on the mechanism which is responsible for the EC growth inhibition. Our model incorporates two possible mechanisms for inhibition. That is, the mechanical cell-cell contact inhibition, and the inhibition mediated from diffusive TGF-b secreted once the two cell types come in contact. Interestingly, our model results suggest that the effective range of TGF-b is a crucial determinant of the degree of EC growth inhibition. Chapter 5, by using a CPM, is devoted to sprouting angiogenesis (the formation of new blood vessel). The dynamic interchange between stalk and tip EC phenotype is incorporated through the Notch signalling pathway, with the leading tip cell moving up macrophage-mediated VEGFA gradients in a non-uniform matrix environment. The model reproduces phenomena in sprouting angiogenesis, including sprout morphology, tip competition, and explains knockout experiments on the Notch signalling pathway. Finally, we close with Chapter 6 where we summarise the ain results from each chapter and propose model extensions for future directions

Integrative modelling of angiogenesis in the bovine corpus luteum

The corpus luteum (CL) is a tissue formed from the remnants of an ovulated follicle in the ovary, and it produces the progesterone needed for a healthy pregnancy. CL growth is highly dependent on a growing nutrient supply, and can be compared with the most aggressive vascular tumours. Angiogenesis, the growth of new blood vessels from existing ones, plays a key role in the growth and function of the CL. Inadequate angiogenesis has been linked to infertility in cows. The CL is composed of several vascular(e.g. endothelial cells (ECs), pericytes (PCs)), and avascular (e.g. luteal cells (LCs), immune cells) cell types, and several pro-angiogenic factors (e.g. Fibroblast Growth Factor 2, FGF2) found to be important in the angiogenic process. The objective of this thesis is to shed light on the cellular and extracellular level determinants of angiogenesis in the bovine CL. We begin with the relevant biological and mathematical literature in Chapter 1. In Chapter 2, an ordinary differential equation model of CL growth is introduced. We assume that the CL volume is a continuum of three cell types, ECs, LCs, and stromal cells (such as PCs). The fourth variable in the model, FGF2, enhances the EC proliferation rate. The model is able, by varying parameters such as the maximal proliferation rate of the ECs, to distinguish cases where the CL shifts from a ‘normal’ to a ‘pathological’ growth. In Chapter 3, we present in vitro CL published and novel studies from Robinson’s Lab. Preliminary results demonstrate interesting endothelial and pericyte behaviours regarding cell aggregation and sprout formation, which are the motivation for the next two Chapters. In these experimental studies, all the CL cell types were incorporated in the same in vitro culture, hence providing a closer approximation to the in vivo environment compared to other in vitro cultures which use only a single cell type (mainly ECs). However, this complicates matters in terms of distinguishing cell behaviours and factors which contribute on the overall cell dynamics. Therefore, in the Chapters 4 and 5 we use data from literature. In Chapter 4, by using the Cellular Potts Model (CPM) framework, we focus on EC-PC interactions, and particularly on the mechanism which is responsible for the EC growth inhibition. Our model incorporates two possible mechanisms for inhibition. That is, the mechanical cell-cell contact inhibition, and the inhibition mediated from diffusive TGF-b secreted once the two cell types come in contact. Interestingly, our model results suggest that the effective range of TGF-b is a crucial determinant of the degree of EC growth inhibition. Chapter 5, by using a CPM, is devoted to sprouting angiogenesis (the formation of new blood vessel). The dynamic interchange between stalk and tip EC phenotype is incorporated through the Notch signalling pathway, with the leading tip cell moving up macrophage-mediated VEGFA gradients in a non-uniform matrix environment. The model reproduces phenomena in sprouting angiogenesis, including sprout morphology, tip competition, and explains knockout experiments on the Notch signalling pathway. Finally, we close with Chapter 6 where we summarise the ain results from each chapter and propose model extensions for future directions

General analysis of breed-and-burn reactors and limited-separations fuel cycles

Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Nuclear Science and Engineering, February 2011.Cataloged from PDF version of thesis. "February 2011."Includes bibliographical references (p. 348-351).A new theoretical framework is introduced, the "neutron excess" concept, which is useful for analyzing breed-and-burn (B&B) reactors and their fuel cycles. Based on this concept, a set of methods has been developed which allows a broad comparison of B&B reactors using different fuels, structural materials, and coolants. This new approach allows important reactor and fuelcycle parameters to be approximated quickly, without the need for a full core design, including minimum burnup/irradiation damage and reactor fleet doubling time. Two general configurations of B&B reactors are considered: a "minimum-burnup" version in which fuel elements can be shuffled in three dimensions, and a "linear-assembly" version composed of conventional linear assemblies that are shuffled radially. Based on studies of different core compositions, the best options for minimizing fuel burnup and material DPA are metal fuel (with a strong dependence on alloy content), the type of steel that allows the lowest structure volume fraction, and helium coolant. If sufficient fuel performance margin exists, sodium coolant can be substituted in place of helium to achieve higher power densities at a modest burnup and DPA penalty. For a minimum-burnup B&B reactor, reasonably achievable minimum DPA values are on the order of 250-350 DPA in steel, while axial peaking in a linear-assembly B&B reactor raises minimum DPA to over 450 DPA. By recycling used B&B fuel in a limited-separations (without full actinide separations) fuel cycle, there is potential for sodium-cooled B&B reactors to achieve fleet doubling times of less than one decade, although this result is highly sensitive to the reactor core composition employed as well as thermal hydraulic performance.by Robert C. Petroski.Ph.D

Impact of pesticides on farmer health and the rice environment

Cet ouvrage présente les conséquences indésirables ou involontaires de l'utilisation des pesticides sur les riziculteurs et les environnements rizicoles. Ses quatorze chapitres et cinq annexes comprennent des revues bibliographiques, des résultats d'enquêtes en milieu paysan, et les résultats d'études au champ et d'études expérimentales conduites entre 1989 et 1991 aux Philippines sous la forme d'un projet multidisciplinaire rassemblant des chercheurs de l'IRRI, du NRI (Angleterre), de l'ORSTOM (France) et de l'UPLB (Philippines). Les études bibliographiques portent sur les méthodes d'analyse des effets de l'utilisation des agrochimiques, le comportement des pesticides appliqués dans les environnements rizicoles, les effets indésirables ou involontaires sur la qualité des eaux, les poissons et les autres vertébrés, les invertébrés aquatiques et telluriques, et la microflore des rizières et le rôle des législations internationales et nationales dans le marché et l'utilisation des pesticides. Les études expérimentales et les enquêtes concernent les effets environnementaux des pesticides sur les environnements rizicoles des Philippines, les perceptions et les connaissances qui gouvernent l'utilisation et les erreurs d'utilisation des pesticides par les riziculteurs et les effets à long terme, médicaux et économiques, de l'exposition des riziculteurs philippins aux pesticides. L'ouvrage comporte également 250 pages de bibliographie annotées sur ces mêmes sujets. (Résumé d'auteur