Multi-monoubiquitylation controls VASP-mediated actin dynamics.

The actin cytoskeleton performs multiple cellular functions, and as such, actin polymerization must be tightly regulated. We previously demonstrated that reversible, non-degradative ubiquitylation regulates the function of the actin polymerase VASP in developing neurons. However, the underlying mechanism of how ubiquitylation impacts VASP activity was unknown. Here, we show that mimicking multi-monoubiquitylation of VASP at K240 and K286 negatively regulates VASP interactions with actin. Using in vitro biochemical assays, we demonstrate the reduced ability of multi-monoubiquitylated VASP to bind, bundle, and elongate actin filaments. However, multi-monoubiquitylated VASP maintained the ability to bind and protect barbed ends from capping protein. Finally, we demonstrate the electroporation of recombinant multi-monoubiquitylated VASP protein altered cell spreading morphology. Collectively, these results suggest a mechanism in which ubiquitylation controls VASP-mediated actin dynamics.

Cerebellin-1 leads the way.

The cerebellin family of proteins influences synapse formation and function. In this issue of PLOS Biology, Han and colleagues identify a new role for Cerebellin-1 in axon growth and guidance.

The transcription factor Myc controls metabolic reprogramming upon T lymphocyte activation.

To fulfill the bioenergetic and biosynthetic demand of proliferation, T cells reprogram their metabolic pathways from fatty acid ß-oxidation and pyruvate oxidation via the TCA cycle to the glycolytic, pentose-phosphate, and glutaminolytic pathways. Two of the top-ranked candidate transcription factors potentially responsible for the activation-induced T cell metabolic transcriptome, HIF1α and Myc, were induced upon T cell activation, but only the acute deletion of Myc markedly inhibited activation-induced glycolysis and glutaminolysis in T cells. Glutamine deprivation compromised activation-induced T cell growth and proliferation, and this was partially replaced by nucleotides and polyamines, implicating glutamine as an important source for biosynthetic precursors in active T cells. Metabolic tracer analysis revealed a Myc-dependent metabolic pathway linking glutaminolysis to the biosynthesis of polyamines. Therefore, a Myc-dependent global metabolic transcriptome drives metabolic reprogramming in activated, primary T lymphocytes. This may represent a general mechanism for metabolic reprogramming under patho-physiological conditions.

Relativistic Effects on a Metal-Metal Bond: Osmium Corrole Dimers.

A series of metal-metal bonded osmium corrole dimers, {Os[T pXPC]}2, were synthesized in reasonably good yields (35-46%) via the interaction of the corresponding free-base meso-tris( p-X-phenyl)corroles (H3[T pXPC], X = CF3, H, CH3, and OCH3), Os3(CO)12, and potassium carbonate in 1,2,4-trichlorobenzene under an inert atmosphere at 180 °C over several hours. The complexes are only the second class of Os corroles reported to date (the first being OsVIN corroles) and also the second class of metal-metal bonded metallocorrole dimers (the other being Ru corrole dimers). Comparison of the X-ray structures, redox potentials, and optical spectra of analogous Ru and Os corrole dimers, along with scalar-relativistic DFT calculations, has provided an experimentally calibrated account of relativistic effects in these complexes. Three of the Os corrole dimers (X = CF3, H, and OCH3) were analyzed with single-crystal X-ray diffraction analysis, revealing inversion-related corrole rings with eclipsed Os-N bonds and Os-Os distances of ∼2.24 Å that are ∼0.06 Å longer than the Ru-Ru distances in the analogous Ru corrole dimers. Interestingly, a comparison of scalar-relativistic and nonrelativistic DFT calculations indicates that this difference in metal-metal bond distance does not, in fact, reflect a differential relativistic effect. For a given corrole ligand, the Ru and Os corrole dimers exhibit nearly identical oxidation potentials but dramatically different reduction potentials, with the Os values ∼0.5 V lower relative to Ru, suggesting that whereas oxidation occurs in a ligand-centered manner, reduction is substantially metal-centered, which indeed was confirmed by scalar-relativistic calculations. The calculations further indicate that approximately a third of the ∼0.5 V difference in reduction potentials can be ascribed to relativity. The somewhat muted value of this relativistic effect appears to be related to the finding that reduction of an Os corrole dimer is not exclusively metal-based but that a significant amount of spin density is delocalized over to the corrole ligand; in contrast, reduction of an Ru corrole dimer occurs exclusively on the Ru-Ru linkage. For isoelectronic complexes, the Ru and Os corrole dimers exhibit substantially different UV-vis spectra. A key difference is a strong near-UV feature of the Os series, which in energy terms is blue-shifted by ∼0.55 V relative to the analogous feature of the Ru series. TDDFT calculations suggest that this difference may be related to higher-energy Os(5d)-based LUMOs in the Os case relative to analogous MOs for Ru.

Rapid one-pot synthesis of pyrrole-appended isocorroles.

Free-base meso-triarylcorroles have been found to undergo oxidative coupling with an excess of pyrrole in dichloromethane in the presence of 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ) affording 5/10-pyrrole-appended isocorroles in reasonable yields (35-60%) and in a matter of seconds. The free-base isocorrole ligands could all be complexed to copper with Cu(OAc)2·H2O in chloroform/methanol in 55-80% yields. Single-crystal X-ray structures of two of the new compounds (H2[5-pyr-TpOMePiC] and Cu[10-pyr-TpOMePiC]) revealed planar macrocycles with rms atomic displacements of only 0.02 and 0.06 Å relative to their respective best-fit C19N4 planes. Both free-base and Cu(ii)-complexed isocorroles exhibit richly featured UV-vis-NIR spectra with red/NIR absorption maxima at ∼650 nm and ∼725 nm for the free-bases and ∼800-850 nm for the copper complexes, suggesting potential applications in photodynamic therapy. Cyclic voltammetric analyses of five of the Cu complexes revealed fully reversible redox cycles with multiple oxidation and reduction features.

A unified model of mammalian BCL-2 protein family interactions at the mitochondria.

During apoptosis, the BCL-2 protein family controls mitochondrial outer membrane permeabilization (MOMP), but the dynamics of this regulation remain controversial. We employed chimeric proteins composed of exogenous BH3 domains inserted into a tBID backbone that can activate the proapoptotic effectors BAX and BAK to permeabilize membranes without being universally sequestered by all antiapoptotic BCL-2 proteins. We thus identified two "modes" whereby prosurvival BCL-2 proteins can block MOMP, by sequestering direct-activator BH3-only proteins ("MODE 1") or by binding active BAX and BAK ("MODE 2"). Notably, we found that MODE 1 sequestration is less efficient and more easily derepressed to promote MOMP than MODE 2. Further, MODE 2 sequestration prevents mitochondrial fusion. We provide a unified model of BCL-2 family function that helps to explain otherwise paradoxical observations relating to MOMP, apoptosis, and mitochondrial dynamics.

A Three-Dimensional Dynamic Supramolecular "Sticky Fingers" Organic Framework.

Engineering high-recognition host-guest materials is a burgeoning area in basic and applied research. The challenge of exploring novel porous materials with advanced functionalities prompted us to develop dynamic crystalline structures promoted by soft interactions. The first example of a pure molecular dynamic crystalline framework is demonstrated, which is held together by means of weak "sticky fingers" van der Waals interactions. The presented organic-fullerene-based material exhibits a non-porous dynamic crystalline structure capable of undergoing single-crystal-to-single-crystal reactions. Exposure to hydrazine vapors induces structural and chemical changes that manifest as toposelective hydrogenation of alternating rings on the surface of the [60]fullerene. Control experiments confirm that the same reaction does not occur when performed in solution. Easy-to-detect changes in the macroscopic properties of the sample suggest utility as molecular sensors or energy-storage materials.

Halterman Corroles and Their Use as a Probe of the Conformational Dynamics of the Inherently Chiral Copper Corrole Chromophore.

Halterman corroles have been synthesized for the first time from pyrrole and Halterman's aldehyde via Gryko's "water-methanol method". These were derivatized to the corresponding copper complexes and subsequently to the ß-octabromo complexes. Electronic circular dichroism spectra were recorded for the enantiopure copper complexes, affording the first such measurements for the inherently chiral Cu corrole chromophore. Interestingly, for a given configuration of the Halterman substituents, X-ray crystallographic studies revealed both P and M conformations of the Cu-corrole core, proving that the substituents, even in conjunction with ß-octabromination, are unable to lock the Cu-corrole core into a given chirality. The overall body of evidence strongly indicates a dynamic equilibrium between the P and M conformations. Such an interconversion, which presumably proceeds via saddling inversion, provides a rationale for our failure so far to resolve sterically hindered Cu corroles into their constituent enantiomers by means of chiral HPLC.

Electronic Structure of Manganese Corroles Revisited: X-ray Structures, Optical and X-ray Absorption Spectroscopies, and Electrochemistry as Probes of Ligand Noninnocence.

Presented herein is a detailed multitechnique investigation of ligand noninnocence in S = 3/2 manganese corrole derivatives at the formal MnIV oxidation state. The Soret maxima of Mn[T pXPC]Cl (T pXPC = meso-tris( p-X-phenyl)corrole, where X = CF3, H, Me, and OMe) were found to red-shift over a range of 37 nm with increasing electron-donating character of X. For Mn[T pXPC]Ph, in contrast, the complex Soret envelopes were found to be largely independent of X. These observations suggested a noninnocent corrole•2--like ligand for the MnCl complexes and an innocent corrole3- ligand for the MnPh complexes. Single-crystal X-ray structures of three Mn[T pXPC]Cl complexes revealed skeletal bond-length alternations indicative of a noninnocent corrole, while no such alternation was observed for Mn[T pOMePC]Ph. B3LYP density functional theory (DFT) calculations on Mn[TPC]Cl yielded strong spatial separation of the α and ß spin densities, consistent with an antiferromagnetically coupled MnIII-corrole•2- description. By comparison, relatively little spatial separation of the α and ß spin densities was found for Mn[TPC]Ph, consistent with an essentially MnIV-corrole3- description. X-ray absorption of near-edge spectroscopy (XANES) revealed a moderate blue shift of 0.6 eV for the Mn K-pre-edge of Mn[T pCF3PC]Ph and a striking enhancement of the pre-edge intensity, relative to Mn[T pCF3PC]Cl, consistent with a more oxidized, i.e., MnIV, center in Mn[T pCF3PC]Ph. Time-dependent DFT calculations indicated that the enhanced intensity of the Mn K-pre-edge of Mn[T pCF3PC]Ph results from the extra 3d z2 hole, which mixes strongly with the Mn 4p z orbital. Combined with similar results on Fe[TPC]Cl and Fe[TPC]Ph, the present study underscores the considerable potential of metal K-edge XANES in probing ligand noninnocence in first-row transition-metal corroles. Cyclic voltammetry measurements revealed highly negative first reduction potentials for the Mn[T pXPC]Ph series (∼-0.95 V) as well as large electrochemical HOMO-LUMO gaps of ∼1.7 V. The first reductions, however, are irreversible, suggesting cleavage of the Mn-Ph bond.

Enhancement of CO2 Uptake and Selectivity in a Metal-Organic Framework by the Incorporation of Thiophene Functionality.

The complex [Zn2(tdc)2dabco] (H2tdc = thiophene-2,5-dicarboxylic acid; dabco = 1,4-diazabicyclooctane) shows a remarkable increase in carbon dioxide (CO2) uptake and CO2/dinitrogen (N2) selectivity compared to the nonthiophene analogue [Zn2(bdc)2dabco] (H2bdc = benzene-1,4-dicarboxylic acid; terephthalic acid). CO2 adsorption at 1 bar for [Zn2(tdc)2dabco] is 67.4 cm3·g-1 (13.2 wt %) at 298 K and 153 cm3·g-1 (30.0 wt %) at 273 K. For [Zn2(bdc)2dabco], the equivalent values are 46 cm3·g-1 (9.0 wt %) and 122 cm3·g-1 (23.9 wt %), respectively. The isosteric heat of adsorption for CO2 in [Zn2(tdc)2dabco] at zero coverage is low (23.65 kJ·mol-1), ensuring facile regeneration of the porous material. Enhancement by the thiophene group on the separation of CO2/N2 gas mixtures has been confirmed by both ideal adsorbate solution theory calculations and dynamic breakthrough experiments. The preferred binding sites of adsorbed CO2 in [Zn2(tdc)2dabco] have been unambiguously determined by in situ single-crystal diffraction studies on CO2-loaded [Zn2(tdc)2dabco], coupled with quantum-chemical calculations. These studies unveil the role of the thiophene moieties in the specific CO2 binding via an induced dipole interaction between CO2 and the sulfur center, confirming that an enhanced CO2 capacity in [Zn2(tdc)2dabco] is achieved without the presence of open metal sites. The experimental data and theoretical insight suggest a viable strategy for improvement of the adsorption properties of already known materials through the incorporation of sulfur-based heterocycles within their porous structures.

Cobalt- and Rhodium-Corrole-Triphenylphosphine Complexes Revisited: The Question of a Noninnocent Corrole.

A reinvestigation of cobalt-corrole-triphenylphosphine complexes has yielded an unexpectedly subtle picture of their electronic structures. UV-vis absorption spectroscopy, skeletal bond length alternations observed in X-ray structures, and broken-symmetry DFT (B3LYP) calculations suggest partial CoII-corrole•2- character for these complexes. The same probes applied to the analogous rhodium corroles evince no evidence of a noninnocent corrole. X-ray absorption spectroscopic studies showed that the Co K rising edge of Co[TPC](PPh3) (TPC = triphenylcorrole) is red-shifted by ∼1.8 eV relative to the bona fide Co(III) complexes Co[TPC](py)2 and Co[TPP](py)Cl (TPP = tetraphenylporphyrin, py = pyridine), consistent with a partial CoII-corrole•2- description for Co[TPC](PPh3). Electrochemical measurements have shown that both the Co and Rh complexes undergo two reversible oxidations and one to two irreversible reductions. In particular, the first reduction of the Rh corroles occurs at significantly more negative potentials than that of the Co corroles, reflecting significantly higher stability of the Rh(III) state relative to Co(III). Together, the results presented herein suggest that cobalt-corrole-triphenylphosphine complexes are significantly noninnocent with moderate CoII-corrole•2- character, underscoring-yet again-the ubiquity of ligand noninnocence among first-row transition metal corroles.

Structural Diversities in Heterometallic Mn-Ca Cluster Chemistry from the Use of Salicylhydroxamic Acid: {MnIII4Ca2}, {MnII/III6Ca2}, {MnIII/IV8Ca}, and {MnIII8Ca2} Complexes with Relevance to Both High- and Low-Valent States of the Oxygen-Evolving Complex.

One-pot reactions between the [Mn3O(O2CPh)6(py)x]+/0 triangular precursors and either CaBr2·xH2O or CaCl2·6H2O, in the presence of salicylhydroxamic acid (shaH2), have afforded the heterometallic complexes [MnIII4Ca2(O2CPh)4(shi)4(H2O)3(Me2CO)] (1) and (pyH)[MnII2MnIII4Ca2Cl2(O2CPh)7(shi)4(py)4] (2), respectively, in good yields. Further reactions but using a more flexible synthetic scheme comprising the Mn(NO3)2·4H2O/Ca(NO3)2·4H2O and Mn(O2CPh)2·2H2O/Ca(ClO4)2·4H2O "metal blends" and shaH2, in the presence of external base NEt3, led to the new complexes (NHEt3)2[MnIII4MnIV4Ca(OEt)2(shi)10(EtOH)2] (3) and (NHEt3)4[MnIII8Ca2(CO3)4(shi)8] (4), respectively. In all reported compounds, the anion of the tetradentate (N,O,O,O)-chelating/bridging ligand salicylhydroxime (shi3-), resulting from the in situ metal-ion-assisted amide-iminol tautomerism of shaH2, was found to bridge both Mn and Ca atoms. Complexes 1-4 exhibit a variety of different structures, metal stoichiometries, and Mn oxidation-state descriptions; 1 possesses an overall octahedral metal arrangement, 2 can be described as a Mn4Ca2 octahedron bound to an additional Mn2 unit, 3 consists of a Mn8 "ring" surrounding a CaII atom, and 4 adopts a rectangular cuboidal motif of eight Mn atoms accommodating two CaII atoms. Solid-state direct-current magnetic susceptibility studies revealed the presence of predominant antiferromagnetic exchange interactions between the Mn centers, leading to S = 0 spin ground-state values for all complexes. From a bioinorganic chemistry perspective, the reported compounds may demonstrate some relevance to both high-valent scheme (3) and lower-oxidation-level species (1, 2, and 4) of the catalytic cycle of the oxygen-evolving complex.

Electronic Structure of Cobalt-Corrole-Pyridine Complexes: Noninnocent Five-Coordinate Co(II) Corrole-Radical States.

Two sets of complexes of Co-triarylcorrole-bispyridine complexes, Co[TpXPC](py)2 and Co[Br8TpXPC](py)2 have been synthesized, where TpXPC refers to a meso-tris(para-X-phenyl)corrole ligand with X = CF3, H, Me, and OMe and Br8TpXPC to the corresponding ß-octabrominated ligand. The axial pyridines in these complexes were found to be labile and, in dilute solutions in dichloromethane, the complexes dissociate almost completely to the five-coordinate monopyridine complexes. Upon addition of a small quantity of pyridine, the complexes revert back to the six-coordinate forms. These transformations are accompanied by dramatic changes in color and optical spectra. 1H NMR spectroscopy and X-ray crystallography have confirmed that the bispyridine complexes are authentic low-spin Co(III) species. Strong substituent effects on the Soret maxima and broken-symmetry DFT calculations, however, indicate a CoII-corrole•2- formulation for the five-coordinate Co[TpXPC](py) series. The calculations implicate a Co(dz2)-corrole("a2u") orbital interaction as responsible for the metal-ligand antiferromagnetic coupling that leads to the open-shell singlet ground state of these species. Furthermore, the calculations predict two low-energy S = 1 intermediate-spin Co(III) states, a scenario that we have been able to experimentally corroborate with temperature-dependent EPR studies. Our findings add to the growing body of evidence for noninnocent electronic structures among first-row transition metal corrole derivatives.

Catalytic activity of the caspase-8-FLIP(L) complex inhibits RIPK3-dependent necrosis.

Caspase-8 has two opposing biological functions--it promotes cell death by triggering the extrinsic pathway of apoptosis, but also has a survival activity, as it is required for embryonic development, T-lymphocyte activation, and resistance to necrosis induced by tumour necrosis factor-α (TNF-α) and related family ligands. Here we show that development of caspase-8-deficient mice is completely rescued by ablation of receptor interacting protein kinase-3 (RIPK3). Adult animals lacking both caspase-8 and RIPK3 display a progressive lymphoaccumulative disease resembling that seen with defects in CD95 or CD95-ligand (also known as FAS and FASLG, respectively), and resist the lethal effects of CD95 ligation in vivo. We have found that caspase-8 prevents RIPK3-dependent necrosis without inducing apoptosis by functioning in a proteolytically active complex with FLICE-like inhibitory protein long (FLIP(L), also known as CFLAR), and this complex is required for the protective function.

Stabilization and Structure of the cis†Tautomer of a Free-Base Porphyrin.

Single-crystal X-ray analysis of the ß-heptakis(trifluoromethyl)-meso-tetrakis(p-fluorophenyl)porphyrin, H2 [(CF3 )7 TpFPP], has revealed the first example of a stable cis tautomer of a free-base porphyrin, the long-postulated intermediate of porphyrin tautomerism. The stability of the unique molecule appears to reflect a dual origin: a strongly saddled porphyrin skeleton, which alleviates electrostatic repulsion between the two NH protons, and two polarization-enhanced, transannular N-H⋅⋅⋅O-H⋅⋅⋅N hydrogen bond chains, each involving a molecule of water. DFT calculations suggest that the observed tautomer has a lower energy than the alternative, doubly hydrated trans tautomer by some 8.3 kcal mol-1 . A fascinating prospect thus exists that H2 [(CF3 )7 TpFPP]⋅2 H2 O and cognate structures may act as supramolecular synthons, which, given their chirality, may even be amenable to resolution into optically pure enantiomers.

Coordination polymers of 5-substituted isophthalic acid.

The synthesis and characterisation of five coordination polymers - Ni2(mip)2(H2O)8·2H2O (1), Zn6(mip)5(OH)2(H2O)4·7.4H2O (2), Zn6(mip)5(OH)2(H2O)2·4H2O (3), Mn(HMeOip)2 (4), and Mn3(tbip)2(Htbip)2(EtOH)2 (5) - are reported. Preliminary nitric oxide release data on compounds 2 and 3 are also given.

Crystal structure resolution of two different chlorhexidine salts.

Two salts of the chlorhexidine di-cation (H2CHx2+) - (H2CHx)(SO4)·3H2O and (H2CHx)(CO3)·4H2O - have been synthesised and characterised crystallographically.

Liquid-phase enantioselective chromatographic resolution using interpenetrated, homochiral framework materials.

Effective separation of mixtures of enantiomers is of continuing interest in analytical and preparative chromatography, with new materials frequently designed and tested. We report two new enantiomerically pure 2D→3D interpenetrated materials used as stationary liquid chromatographic (LC) phases that are shown to resolve selected racemic mixtures with enantiomeric and chemical selectivity. Dicarboxylate ligands derived from amino acids on naphthalene and perylene cores form 2D frameworks that interpenetrate to give 3D structures. Selectivity is initially tested by uptake from solution; subsequent LC methods show that the materials exhibit resolution of racemic analytes in 'micro-columns' and that the two closely related materials show markedly different selectivity for different analytes with much greater activity than the ligands alone. Comparison with a close-packed analogue suggests that the separation activity is largely due to surface effects.

Synthesis, structure and cation-binding properties of some [4 + 4] metallocyclic MO2(2+) (M = Mo or W) derivatives of 9-phenyl-2,3,7-trihydroxyfluor-6-one.

The trianion Z(3-) obtained from 9-phenyl 2,3,7-trihydroxyfluor-6-one, ZH3, affords dioxomolybdenum and dioxotungsten derivatives which contain [4 + 4] metallocycles of composition [(MO2)4Z4](4-) (M = Mo, W) in combination with a variety of counter cations. The syntheses, structures and ESMS of the following compounds are presented: compound 1, (MePPh3)3(NBu4)[(MoO2)4Z4]; compound 2, (MePPh3)3(NBu4)[(WO2)4Z4]; compound 3, (MePPh3)4[(WO2)4Z4]; compound 4, (PPh4)2(NBu4)2[(MoO2)4Z4]; compound 5, (AsPh4)3(NBu4)[(MoO2)4Z4]; compound 6, (AsPh4)2(NBu4)2[(WO2)4Z4]; compound 7, (Ph3PNPPh3)(NBu4)3[(MoO2)4Z4]; compound 8, (Ph3PNPPh3)(NBu4)3[(WO2)4Z4]; compound 9, (NEt4)(NBu4)3[(MoO2)4Z4]. The metallocycles in all of these compounds have similar structures, with the four metal centers located at the corners of a square slightly distorted, to varying degrees, toward a rhombus and also toward a tetrahedron. Various cations are bound inside the anionic metallocycles. ESI mass spectrometry shows that the metallocycles remain intact in the gas phase, forming [(MO2)4Z4](4-), {X-[(MO2)4Z4]}(3-) and in some cases {X2-[(MO2)4Z4]}(2-) where X(+) is an organic cation.

Loss of the E3 ubiquitin ligase TRIM67 alters the post-synaptic density proteome.

The E3 ubiquitin ligase TRIM67 is enriched in the central nervous system and is required for proper neuronal development. Previously we demonstrated TRIM67 coordinates with the closely related E3 ubiquitin ligase TRIM9 to regulate cytoskeletal dynamics downstream of the netrin-1 during axon guidance and axon branching in early neuronal morphogenesis. Interestingly, loss of Trim67 impacts cognitive flexibility in a spatial learning and memory task. Despite this behavioral phenotype, it was previously uninvestigated if TRIM67 was involved in synapse formation or function. Here we demonstrate TRIM67 localizes to the post-synaptic density (PSD) within dendritic spines. Furthermore, we show that loss of Trim67 significantly changes a subset of proteins within the PSD proteome, including changes in the regulation of the actin and microtubule cytoskeletons. Collectively, our data propose a synaptic role for TRIM67.