A Toolkit of Engineered Recombinational Balancers in C. elegans

Dejima and colleagues report using CRISPR/Cas9 to generate a new collection of greatly improved balancer chromosomes in the standard laboratory nematode Caenorhabditis elegans, using methods previously reported by the same laboratory, expanding the set of C. elegans balancers to cover nearly 90% of coding genes

The genetics of the sexually dimorphic deaths of the C. elegans CEM neutrons

Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Biology, 2009.Includes bibliographical references.The cells of metazoan organisms possess the capability to commit a form of cellular suicide known as programmed cell death or apoptosis. The proper control of this endogenous death program is essential to animal development and to the prevention of disease. To better understand how individual cells are developmentally specified to die, I studied the survival decision of a single cell type in the nematode Caenorhabditis elegans, the CEM neurons. The CEMs die during hermaphrodite embryogenesis and survive and function as sensory neurons in males. I identified 144 independent mutant strains in which the CEM neurons of hermaphrodites survive, including 52 mutants in known cell-death genes and 67 mutants generally defective sex determination. Another 29 screen isolates displayed a new defect of transcriptional de repression, the green pharynx phenotype. From these isolates and from additional screens, I defined a set of seven genes that function to prevent inappropriate gene expression. From the isolates causing CEM survival I identified two new sex-determination proteins, the PLZF-like transcription factor TRA-4 and the F-box protein SEL-10; demonstrated that the neurogenesis genes vab-3 Pax6 and cnd-1 NeuroD are required for aspects of the CEM fate, including CEM death, and likely function together in this process and in other aspects of C. elegans development, a cooperative relationship likely to be evolutionarily conserved; and identified the Bar family homeodomain transcription factor gene ceh-30 as specifically promoting CEM survival. The CEM neurons of males lacking ceh-30 inappropriately undergo programmed cell death. In the CEMs of hermaphrodites, ceh-30 is directly repressed by TRA-1, a transcription factor that acts as the last step in the sex-determination pathway to promote a hermaphrodite identity.(cont.) The cell-protective function of ceh-30 is specific to the CEM neurons, and ceh-30 is expressed in and acts cell-autonomously in the CEMs to promote their survival. Expression of the mouse ceh-30 homolog Barhll can restore CEM survival to ceh-30 mutants. In mice lacking Barhl1, as in ceh-30 mutants, a specific class of sensory neuron is generated normally differentiates, but subsequently inappropriately undergoes apoptotic cell death. Protection of the CEM neurons by ceh-30 does not require CED-9, the sole member of the multidomain Bcl-2 family in C. elegans. By contrast, other regulators of the survival decisions of specific cells in C. elegans act through transcriptional control of the CED-9 inhibitor egl-1. Mammalian regulation of cell death is similarly almost entirely mediated through members of the multidomain Bcl-2 family. The evolutionarily conserved cell-protective function of ceh-30 therefore probably defines a previously unknown mechanism capable of promoting cell survival both in nematode development, in the sensory neurons required for hearing in the mouse and likely in humans.by Hillel Tsvi Schwartz.Ph.D

Fault Tolerant Max-Cut

In this work, we initiate the study of fault tolerant Max-Cut, where given an edge-weighted undirected graph G = (V,E), the goal is to find a cut S ? V that maximizes the total weight of edges that cross S even after an adversary removes k vertices from G. We consider two types of adversaries: an adaptive adversary that sees the outcome of the random coin tosses used by the algorithm, and an oblivious adversary that does not. For any constant number of failures k we present an approximation of (0.878-?) against an adaptive adversary and of ?_{GW}? 0.8786 against an oblivious adversary (here ?_{GW} is the approximation achieved by the random hyperplane algorithm of [Goemans-Williamson J. ACM `95]). Additionally, we present a hardness of approximation of ?_{GW} against both types of adversaries, rendering our results (virtually) tight. The non-linear nature of the fault tolerant objective makes the design and analysis of algorithms harder when compared to the classic Max-Cut. Hence, we employ approaches ranging from multi-objective optimization to LP duality and the ellipsoid algorithm to obtain our results

Enhanced bioavailability and reduced pharmacokinetic variability of Oral PTH (1-34) in man

An orally administered PTH may have prodigious advantages in the treatment of hypoparathyroidism and osteoporosis. Unfortunately, the oral delivery of biologic macromolecules is characterized by a negligible bioavailability and a high dose-to-dose variability in absorption, resulting in difficulty in accurately titrating the drug effect. We present clinical study data of a novel oral peptide delivery technology demonstrating an enhanced bioavailability with reduced Cmax variability. Methods: A Phase I, open label crossover pharmacokinetic (PK) study to assess the safety and PK of oral PTH (1-34) in ten healthy male adult volunteers was conducted. The PK profile of a fixed dose - 1.5mg PTH (1-34) of three different oral formulations was compared. PTH (1-34) levels in the plasma of subjects was analyzed at a number of time points post administration, utilizing a PTH (1-34) immunoassay (IDS; Bolden, UK). In parallel, to assess the pharmacodynamic (PD) effect, serum calcium of subjects receiving the different formulations of oral PTH (1-34) was analyzed. Results: PK profiles of all oral PTH (1-34) formulations were characterized by a rapid absorption and elimination. The systemic exposure (AUC) of the basic oral formulation and two modified formulation versions were 3481 ±1843 pg*min/mL, 7976 ±2556 pg*min/mL and 11369 ±3719 pg*min/mL (mean ± SE). The maximal plasma concentration (Cmax) of these formulations were 145 ±56pg/mL, 375 ±108pg/mL, and 481 ±101pg/mL, respectively. Cmax coefficients of variation (CV%) of the same formulations were 123%, 91% and 67%, respectively. Similarly to the drug absorption, PD response of the modified formulations, presented as the maximal relative increase in albumin adjusted calcium, was improved from 0.07 ±0.29mg/dL to 0.32 ±0.24mg/dL. Discussion: Inherent to oral drug delivery of biopharmaceuticals is the extremely low bioavailability and high absorption variability. The current results indicate that Entera’s delivery technology can overcome these two principal obstacles by achieving repeatable, clinically relevant systemic drug exposure. Entera’s proprietary delivery platform was optimized and achieved anenhancement in drug bioavailability in parallel with the significant decrease in its absorption variability. Similarly, its effect on blood calcium was enhanced by the novel oral formulation of PTH (1-34) pointing out the potential of the drug to be a first line treatment of hypoparathyroidism and osteoporosis

Il Rumore Lontano

Il 20 febbraio 1898, la popolazione svizzera approvò con 386’634 voti favorevoli e 182’718 contrari la «Legge federale sull’acquisto e la gestione di ferrovie per conto della Confederazione e l’organizzazione amministrativa delle Ferrovie Federali Svizzere». Il Capodanno del 1901, alle ore 2.00, alla stazione di Berna si festeggiò l’arrivo del primo treno proveniente da Zurigo–Aarau–Olten, che poi proseguì per Losanna–Ginevra. A questa rivoluzione se ne aggiunge un’altra. Nel 1914 l’ingegnere inglese Richard Trevithick presenta in pubblico il primo prototipo di locomotiva per il trasporto ferroviario. Da quel giorno il treno veloce si è diffuso in tutto il mondo ed è diventato il primo vero e proprio veicolo di trasporto di massa. Ha rappresentato inoltre un punto di svolta per l’evoluzione industriale delle nazioni del novecento, arrivando a rivestire un ruolo centrale nella struttura politica, economica e sociale delle nazioni. L’invenzione della locomotiva ha aperto quindi una nuova era della storia dell’umanità rendendo accessibile il trasporto di merci e persone su brevi e lunghe distanze e cambiando per sempre la percezione dello spazio e del tempo. Grazie al treno il mondo non è stato più lo stesso. In un paio di secoli la locomotiva e in generale il treno con tutte le connotazioni che porta con sé (il viaggio, il fischio, il classico rumore,…) è diventato parte dell’immaginario collettivo e, nelle sue molteplici variazioni e sviluppi tecnologici, un soundmark geografico ovvero un segno identitario. In Svizzera, ed in particolare in Canton Ticino il territorio oggetto di questo studio, la complessa orografia ha sviluppato un’ingegneria ferroviaria storicamente all’avanguardia in ambito internazionale

Thermal Stability of the Human Immunodeficiency Virus Type 1 (HIV-1) Receptors, CD4 and CXCR4, Reconstituted in Proteoliposomes

Background: The entry of human immunodeficiency virus (HIV-1) into host cells involves the interaction of the viral exterior envelope glycoprotein, gp120, and receptors on the target cell. The HIV-1 receptors are CD4 and one of two chemokine receptors, CCR5 or CXCR4. Methodology/Principal Findings: We created proteoliposomes that contain CD4, the primary HIV-1 receptor, and one of the coreceptors, CXCR4. Antibodies against CD4 and CXCR4 specifically bound the proteoliposomes. CXCL12, the natural ligand for CXCR4, and the small-molecule CXCR4 antagonist, AMD3100, bound the proteoliposomes with affinities close to those associated with the binding of these molecules to cells expressing CXCR4 and CD4. The HIV-1 gp120 exterior envelope glycoprotein bound tightly to proteoliposomes expressing only CD4 and, in the presence of soluble CD4, bound weakly to proteoliposomes expressing only CXCR4. The thermal stability of CD4 and CXCR4 inserted into liposomes was examined. Thermal denaturation of CXCR4 followed second-order kinetics, with an activation energy (E$_a$) of 269 kJ/mol (64.3 kcal/mol) and an inactivation temperature (T$_i$) of 56°C. Thermal inactivation of CD4 exhibited a reaction order of 1.3, an E$_a$ of 278 kJ/mol (66.5 kcal/mol), and a T$_i$ of 52.2°C. The second-order denaturation kinetics of CXCR4 is unusual among G protein-coupled receptors, and may result from dimeric interactions between CXCR4 molecules. Conclusions/Significance: Our studies with proteoliposomes containing the native HIV-1 receptors allowed an examination of the binding of biologically important ligands and revealed the higher-order denaturation kinetics of these receptors. CD4/CXCR4-proteoliposomes may be useful for the study of virus-target cell interactions and for the identification of inhibitors