A mass spectrometry workflow for measuring protein turnover rates in vivo

Proteins are continually produced and degraded, to avoid the accumulation of old or damaged molecules and to maintain the efficiency of physiological processes. Despite its importance, protein turnover has been difficult to measure in vivo. Previous approaches to evaluating turnover in vivo have required custom labeling approaches, involved complex mass spectrometry (MS) analyses, or used comparative strategies that do not allow direct quantitative measurements. Here, we describe a robust protocol for quantitative proteome turnover analysis in mice that is based on a commercially available diet for stable isotope labeling of amino acids in mammals (SILAM). We start by discussing fundamental concepts of protein turnover, including different methodological approaches. We then cover in detail the practical aspects of metabolic labeling and explain both the experimental and computational steps that must be taken to obtain accurate in vivo results. Finally, we present a simple experimental workflow that enables measurement of precise turnover rates in a time frame of similar to 4-5 weeks, including the labeling time. We also provide all the scripts needed for the interpretation of the MS results and for comparing turnover across different conditions. Overall, the workflow presented here comprises several improvements in the determination of protein lifetimes with respect to other available methods, including a minimally invasive labeling strategy and a robust interpretation of MS results, thus enhancing reproducibility across laboratories

On the Possibility of Accelerating Positron on an Electron Wake at SABER

A new approach for positron acceleration in non-linear plasma wakefields driven by electron beams is presented. Positrons can be produced by colliding an electron beam with a thin foil target embedded in the plasma. Integration of positron production and acceleration in one stage is realized by a single relativistic, intense electron beam. Simulations with the parameters of the proposed SABER facility [1] at SLAC suggest that this concept could be tested there

Ketogenic diet uncovers differential metabolic plasticity of brain cells

To maintain homeostasis, the body, including the brain, reprograms its metabolism in response to altered nutrition or disease. However, the consequences of these challenges for the energy metabolism of the different brain cell types remain unknown. Here, we generated a proteome atlas of the major central nervous system (CNS) cell types from young and adult mice, after feeding the therapeutically relevant low-carbohydrate, high-fat ketogenic diet (KD) and during neuroinflammation. Under steady-state conditions, CNS cell types prefer distinct modes of energy metabolism. Unexpectedly, the comparison with KD revealed distinct cell type–specific strategies to manage the altered availability of energy metabolites. Astrocytes and neurons but not oligodendrocytes demonstrated metabolic plasticity. Moreover, inflammatory demyelinating disease changed the neuronal metabolic signature in a similar direction as KD. Together, these findings highlight the importance of the metabolic cross-talk between CNS cells and between the periphery and the brain to manage altered nutrition and neurological disease

High-gradient plasma and laser accelerators

Novel high-gradient accelerators have demonstrated acceleration of electrons and positrons with electric field strengths of 1 to > 100 GeV/m. This is about 10 to 1000 times higher than achieved in RF-based accelerators, and as such they have the potential to overcome the limitations associated with RF cavities. Plasma-based accelerators have produced multi-GeV bunches with parameters approaching those suitable for a linear collider. These accelerators offer the prospect of near term, compact and cost-effective particle physics experiments that provide new physics possibilities supporting precision studies and the search for new particles. The expert panel has defined a long term R&D roadmap towards a compact collider with attractive intermediate experiments and studies. A delivery plan for the required R&D has been developed and includes work packages, deliverables, a minimal plan, connections to ongoing projects and an aspirational plan

Structure Loaded Vacuum Laser-Driven Particle Acceleration Experiments at SLAC

We present an overview of the future laser-driven particle acceleration experiments. These will be carried out at the E163 facility at SLAC. Our objectives include a reconfirmation of the proof-of-principle experiment, a staged buncher laser-accelerator experiment, and longer-term future experiments that employ dielectric laser-accelerator microstructures

Inverse-Transition Radiation Laser Acceleration Experiments at SLAC

We present a series of laser-driven particle acceleration experiments that are aimed at studying laser-particle acceleration as an inverse-radiation process. To this end we employ a semi-open vacuum setup with a thin planar boundary that interacts with the laser and the electromagnetic field of the electron beam. Particle acceleration from different types of boundaries will be studied and compared to the theoretical expectations from the Inverse-radiation picture and the field path integral method. We plan to measure the particle acceleration effect from transparent, reflective, black, and rough surface boundaries. While the agreement between the two acceleration pictures is straightforward to prove analytically for the transparent and reflective boundaries the equivalence is not clear-cut for the absorbing and rough-surface boundaries. Experimental observation may provide the evidence to distinguish between the models

Beam Coupling to Optical Scale Accelerating Structures

Current research efforts into structure based laser acceleration of electrons utilize beams from standard RF linacs. These beams must be coupled into very small structures with transverse dimensions comparable to the laser wavelength. To obtain decent transmission, a permanent magnet quadrupole (PMQ) triplet with a focusing gradient of 560 T/m is used to focus into the structure. Also of interest is the induced wakefield from the structure, useful for diagnosing potential accelerator structures or as novel radiation sources

Arabidopsis AtVPS15 is essential for pollen development and germination through modulating phosphatidylinositol 3-phosphate formation

Arabidopsis thaliana phosphatidylinositol 3-kinase (AtVPS34) functions in the development and germination of pollen by catalyzing the biosynthesis of phosphatidylinositol 3-phosphate (PI3P). In yeast, Vps15p is required for the membrane targeting and activity of Vps34. The expression of Arabidopsis thaliana VPS15 (AtVPS15), an ortholog of yeast Vps15, is mainly detected in pollen grains and pollen tubes. To determine its role in pollen development and pollen tube growth, we attempted to isolate the T-DNA insertion mutants of AtVPS15; however, homozygous lines of atvps15 were not obtained from the progeny of atvps15/+ heterozygotes. Genetic analysis revealed that the abnormal segregation is due to the failure of transmission of the atvps15 allele through pollen. Most pollen grains from the atvps15/+ genotype are viable, with normal exine structure and nuclei, but some mature pollen grains are characterized with unusual large vacuoles that are not observed in pollen grains from the wild AtVPS15 genotype. The germination ratio of pollen from the atvps15/+ genotype is about half when compared to that from the wild AtVPS15 genotype. When supplied with PI3P, in vitro pollen germination of the atvps15/+ genotype is greatly improved. Presumably, AtVPS15 functions in pollen development and germination by regulating PI3P biosynthesis in Arabidopsis