Improved split fluorescent proteins for endogenous protein labeling.

Self-complementing split fluorescent proteins (FPs) have been widely used for protein labeling, visualization of subcellular protein localization, and detection of cell-cell contact. To expand this toolset, we have developed a screening strategy for the direct engineering of self-complementing split FPs. Via this strategy, we have generated a yellow-green split-mNeonGreen21-10/11 that improves the ratio of complemented signal to the background of FP1-10-expressing cells compared to the commonly used split GFP1-10/11; as well as a 10-fold brighter red-colored split-sfCherry21-10/11. Based on split sfCherry2, we have engineered a photoactivatable variant that enables single-molecule localization-based super-resolution microscopy. We have demonstrated dual-color endogenous protein tagging with sfCherry211 and GFP11, revealing that endoplasmic reticulum translocon complex Sec61B has reduced abundance in certain peripheral tubules. These new split FPs not only offer multiple colors for imaging interaction networks of endogenous proteins, but also hold the potential to provide orthogonal handles for biochemical isolation of native protein complexes.Split fluorescent proteins (FPs) have been widely used to visualise proteins in cells. Here the authors develop a screen for engineering new split FPs, and report a yellow-green split-mNeonGreen2 with reduced background, a red split-sfCherry2 for multicolour labeling, and its photoactivatable variant for super-resolution use

Theory of Metal-Insulator Transition in PrRu4P12 and PrFe4P12

All symmetry allowed couplings between the 4f^2-electron ground state doublet of trivalent praseodymium in PrRu4P12 and PrFe4P12 and displacements of the phosphorus, iron or ruthenium ions are considered. Two types of displacements can change the crystal lattice from body-centred cubic to simple orthorhombic or to simple cubic. The first type lowers the point group symmetry from tetrahedral to orthorhombic, while the second type leaves it unchanged, with corresponding space group reductions Im3 --> Pmmm and Im3 --> Pm3 respectively. In former case, the lower point-group symmetry splits the degeneracy of the 4f^2 doublet into states with opposite quadrupole moment, which then leads to anti-quadrupolar ordering, as in PrFe4P12. Either kind of displacement may conspire with nesting of the Fermi surface to cause the metal-insulator or partial metal-insulator transition observed in PrFe4P12 and PrRu4P12. We investigate this scenario using band-structure calculations, and it is found that displacements of the phosphorus ions in PrRu4P12 (with space group reduction Im3 --> Pm3) open a gap everywhere on the Fermi surface.Comment: 6 page

31P-NMR and muSR Studies of Filled Skutterudite Compound SmFe4P12: Evidence for Heavy Fermion Behavior with Ferromagnetic Ground State

The 31P-NMR (nuclear magnetic resonance) and muSR (muon spin relaxation) measurements on the filled skutterudite system SmFe4P12 have been carried out. The temperature T dependence of the 31P-NMR spectra indicates the existence of the crystalline electric field effect splitting of the Sm3+$ (J = 5/2) multiplet into a ground state and an excited state of about 70 K. The spin-lattice relaxation rate 1/T1 shows the typical behavior of the Kondo system, i.e., 1/T1 is nearly T independent above 30 K, and varies in proportion to T (the Korringa behavior, 1/T1 \propto T) between 7.5 K and 30 K. The T dependence deviated from the Korringa behavior below 7 K, which is independent of T in the applied magnetic field of 1 kOe, and suppressed strongly in higher fields. The behavior is explained as 1/T1is determined by ferromagnetic fluctuations of the uncovered Sm3+ magnetic moments by conduction electrons. The muSR measurements in zero field show the appearance of a static internal field associated with the ferromagnetic order below 1.6 K.Comment: 6 pages, 9 figures, to be published in J. Phys. Soc. Jpn. 75 (2006

XANES study of rare-earth valency in LRu4P12 (L = Ce and Pr)

Valency of Ce and Pr in LRu4P12 (L = Ce and Pr) was studied by L2,3-edge x-ray absorption near-edge structure (XANES) spectroscopy. The Ce-L3 XANES spectrum suggests that Ce is mainly trivalent, but the 4f state strongly hybridizes with ligand orbitals. The band gap of CeRu4P12 seems to be formed by strong hybridization of 4f electrons. Pr-L2 XANES spectra indicate that Pr exists in trivalent state over a wide range in temperature, 20 < T < 300 K. We find that the metal-insulator (MI) transition at TMI = 60 K in PrRu4P12 does not originate from Pr valence fluctuation.Comment: 4 page

Composition of Ices in Low-Mass Extrasolar Planets

We study the formation conditions of icy planetesimals in protoplanetary disks in order to determine the composition of ices in small and cold extrasolar planets. Assuming that ices are formed from hydrates, clathrates, and pure condensates, we calculate their mass fractions with respect to the total quantity of ices included in planetesimals, for a grid of disk models. We find that the composition of ices weakly depends on the adopted disk thermodynamic conditions, and is rather influenced by the initial composition of the gas phase. The use of a plausible range of molecular abundance ratios and the variation of the relative elemental carbon over oxygen ratio in the gas phase of protoplanetary disks, allow us to apply our model to a wide range of planetary systems. Our results can thus be used to constrain the icy/volatile phase composition of cold planets evidenced by microlensing surveys, hypothetical ocean-planets and carbon planets, which could be detected by Corot or Kepler.Comment: Accepted for publication in The Astrophysical Journa

Confirmation of a one-dimensional spin-1/2 Heisenberg system with ferromagnetic first-nearest-neighbor and antiferromagnetic second-nearest-neighbor interactions in Rb2{}_{2}Cu2{}_{2}Mo3{}_{3}O12{}_{12}

We have investigated magnetic properties of Rb$_2$Cu$_2$Mo$_3$O$_{12}$ powder. Temperature dependence of magnetic susceptibility and magnetic-field dependence of magnetization have shown that this cuprate is a model compound of a one-dimensional spin-1/2 Heisenberg system with ferromagnetic first-nearest-neighbor (1NN) and antiferromagnetic second-nearest-neighbor (2NN) competing interactions (competing system). Values of the 1NN and 2NN interactions are estimated as $J_1 = -138$ K and $J_2 = 51$ K ($\alpha \equiv J_2 / J_1 = -0.37$). This value of $\alpha$ suggests that the ground state is a spin-singlet incommensurate state. In spite of relatively large $J_1$ and $J_2$, no magnetic phase transition appears down to 2 K, while an antiferromagnetic transition occurs in other model compounds of the competing system with ferromagnetic 1NN interaction. For that reason, Rb$_2$Cu$_2$Mo$_3$O$_{12}$ is an ideal model compound to study properties of the incommensurate ground state that are unconfirmed experimentally.Comment: 6 pages, 4 figure

Optical Conductivity and Electronic Structure of CeRu4Sb12 under High Pressure

Optical conductivity [s(w)] of Ce-filled skutterudite CeRu4Sb12 has been measured at high pressure to 8 GPa and at low temperature, to probe the pressure evolution of its electronic structures. At ambient pressure, a mid-infrared peak at 0.1 eV was formed in s(w) at low temperature, and the spectral weight below 0.1 eV was strongly suppressed, due to a hybridization of the f electron and conduction electron states. With increasing external pressure, the mid-infrared peak shifts to higher energy, and the spectral weight below the peak was further depleted. The obtained spectral data are analyzed in comparison with band calculation result and other reported physical properties. It is shown that the electronic structure of CeRu4Sb12 becomes similar to that of a narrow-gap semiconductor under external pressure.Comment: 8 pages, 9 figure

Paradoxical Sensitivity to an Integrated Stress Response Blocking Mutation in Vanishing White Matter Cells

The eukaryotic translation initiation factor eIF2B promotes mRNA translation as a guanine nucleotide exchange factor (GEF) for translation initiation factor 2 (eIF2). Endoplasmic reticulum (ER) stress-mediated activation of the kinase PERK and the resultant phosphorylation of eIF2's alpha subunit (eIF2α) attenuates eIF2B GEF activity thereby inducing an integrated stress response (ISR) that defends against protein misfolding in the ER. Mutations in all five subunits of human eIF2B cause an inherited leukoencephalopathy with vanishing white matter (VWM), but the role of the ISR in its pathogenesis remains unclear. Using CRISPR-Cas9 genome editing we introduced the most severe known VWM mutation, EIF2B4$^{A391D}$, into CHO cells. Compared to isogenic wildtype cells, GEF activity of cells with the VWM mutation was impaired and the mutant cells experienced modest enhancement of the ISR. However, despite their enhanced ISR, imposed by the intrinsic defect in eIF2B, disrupting the inhibitory effect of phosphorylated eIF2α on GEF by a contravening EIF2S1/eIF2α$^{S51A}$ mutation that functions upstream of eIF2B, selectively enfeebled both EIF2B4$^{A391D }$ and the related severe VWM EIF2B4$^{R483W}$ cells. The basis for paradoxical dependence of cells with the VWM mutations on an intact eIF2α genotype remains unclear, as both translation rates and survival from stressors that normally activate the ISR were not reproducibly affected by the VWM mutations. Nonetheless, our findings support an additional layer of complexity in the development of VWM, beyond a hyperactive ISR.Supported by grants from the Wellcome Trust (Wellcome 200848/Z/16/Z) and the European Commission (EU FP7 Beta-Bat No: 277713) and, a Wellcome Trust Strategic Award for core facilities to the Cambridge Institute for Medical Research (Wellcome 100140). YS is a Japanese Society for the Promotion of Science Postdoctoral Fellow for Research Abroad. NAW is a Medical Research Council supported PhD student. DR is a Wellcome Trust Principal Research Fellow