A conserved germline-specific Dsn1 alternative splice isoform supports oocyte and embryo development.

Alternative mRNA splicing can generate distinct protein isoforms to allow for the differential control of cell processes across cell types. However, alternative splice isoforms that differentially modulate distinct cell division programs have remained elusive. Here, we demonstrate that mammalian germ cells express an alternate mRNA splice isoform for the kinetochore component, DSN1, a subunit of the MIS12 complex that links the centromeres to spindle microtubules during chromosome segregation.

Optical and EPR Detection of a Triplet Ground State Phenyl Nitrenium Ion.

Nitrenium ions are important reactive intermediates participating in the synthetic chemistry and biological processes. Little is known about triplet phenyl nitrenium ions regarding their reactivity, lifetimes, spectroscopic features, and electronic configurations, and no ground state triplet nitrenium ion has been directly detected. In this work, -pyrrolidinyl-phenyl hydrazine hydrochloride () is synthesized as the photoprecursor to photochemically generate the corresponding -pyrrolidinyl-phenyl nitrenium ion (), which is computed to adopt a π, π* triplet ground state.

Make Selenium Reactive Again: Activating Elemental Selenium for Synthesis of Metal Selenides Ranging from Nanocrystals to Large Single Crystals.

The inertness of elemental selenium is a significant obstacle in the synthesis of selenium-containing materials at low reaction temperatures. Over the years, several recipes have been developed to overcome this hurdle; however, most of the methods are associated with the use of highly toxic, expensive, and environmentally harmful reagents. As such, there is an increasing demand for the design of cheap, stable, and nontoxic reactive selenium precursors usable in the low-temperature synthesis of transition metal selenides with vast applications in nanotechnology, thermoelectrics, and superconductors.

Acidification of colostrum affects the fecal microbiota of preweaning dairy calves.

Calf diarrhea is a leading cause of death in preweaning calves and it causes major economic losses to producers. Acidified milk has been shown to have beneficial effects on health and growth parameters in calves but there is little research into its effects on the microbiota, and few studies on the use of acidified colostrum. The purpose of this study was to compare how feeding acidified colostrum to calves at birth affects fecal microbiota from birth through 8 wk of age compared with calves fed nonacidified colostrum.

Effect of formic acid treatment on colostrum quality, and on absorption and function of immunoglobulins: a randomized controlled trial in Holstein dairy calves.

Background: Good quality colostrum is characterized by high immunoglobulin concentration and low pathogen load. Some methods of pathogen reduction can decrease immunoglobulin concentration and potentially affect their function. Objectives were to determine the effect of formic acid treatment on colostral bacterial and immunoglobulin (IgG) levels before feeding, and serum immunoglobulin concentration and neutralizing capabilities after feeding.

Synthesis-enabled exploration of chiral and polar multivalent quaternary sulfides.

An innovative method of synthesis is reported for the large and diverse (RE)(TM) (Tt)S (RE = rare-earth, TM = transition metals, Tt = Si, Ge, and Sn) family of compounds (∼1000 members, ∼325 contain Si), crystallizing in the noncentrosymmetric, chiral, and polar 6 space group. Traditional synthesis of such phases involves the annealing of elements or binary sulfides at elevated temperatures. The atomic mixing of refractory components technique, presented here, allows the synthesis of known members and vastly expands the family to nearly the entire transition metal block, including 3d, 4d, and 5d TMs with oxidation states ranging from 1+ to 4+.

Effect of exercise on peritoneal microenvironment and progression of ovarian cancer.

Ovarian cancer is one of the deadliest gynecological malignancies and lacks treatments that do not significantly impact patient health-related quality of life. Exercise has been associated with reduced cancer risk and improved clinical outcomes; however the underlying molecular mechanisms are unknown. In this study, we utilized a treadmill-running exercise model to investigate the effects of exercise on high-grade serous ovarian carcinoma (HGSOC) progression and chemotherapy outcomes.

Ovarian BDNF promotes survival, migration, and attachment of tumor precursors originated from p53 mutant fallopian tube epithelial cells.

High-grade serous ovarian carcinoma (HGSOC) is the most lethal gynecological malignancy. New evidence supports a hypothesis that HGSOC can originate from fallopian tube epithelium (FTE). It is unclear how genetic alterations and pathophysiological processes drive the progression of FTE tumor precursors into widespread HGSOCs.

Inhibition of Heat Shock Protein 90 suppresses TWIST1 Transcription.

Molecular chaperone heat shock protein 90 (HSP90) is involved in oncogenic signaling pathways including epithelial-mesenchymal transition (EMT), a key process in tumor initiation, progression, metastasis, and chemoresistance. The molecular mechanisms underlying the involvement of HSP90 in EMT are still under investigation. In this study, we identified a previously unrecognized role of HSP90 in cooperating with signal transducer and activator of transcription 3 (STAT3) to regulate TWIST1 transcription in cancer cells.

Quantitative investigation of free radicals in bio-oil and their potential role in condensed-phase polymerization.

We report on the quantitative analysis of free radicals in bio-oils produced from pyrolysis of cellulose, organosolv lignin, and corn stover by EPR spectroscopy. Also, we investigated their potential role in condensed-phase polymerization. Bio-oils produced from lignin and cellulose show clear evidence of homolytic cleavage reactions during pyrolysis that produce free radicals.

NMR determination of protein partitioning into membrane domains with different curvatures and application to the influenza M2 peptide.

The M2 protein of the influenza A virus acts both as a drug-sensitive proton channel and mediates virus budding through membrane scission. The segment responsible for causing membrane curvature is an amphipathic helix in the cytoplasmic domain of the protein. Here, we use (31)P and (13)C solid-state NMR to examine M2-induced membrane curvature.

Membrane-dependent effects of a cytoplasmic helix on the structure and drug binding of the influenza virus M2 protein.

The influenza A M2 protein forms a proton channel for virus infection and also mediates virus assembly and budding. The minimum protein length that encodes both functions contains the transmembrane (TM) domain (roughly residues 22-46) for the amantadine-sensitive proton-channel activity and an amphipathic cytoplasmic helix (roughly residues 45-62) for curvature induction and virus budding. However, structural studies involving the TM domain with or without the amphipathic helix differed on the drug-binding site.

Specific binding of adamantane drugs and direction of their polar amines in the pore of the influenza M2 transmembrane domain in lipid bilayers and dodecylphosphocholine micelles determined by NMR spectroscopy.

The transmembrane domain of the influenza M2 protein (M2TM) forms a tetrameric proton channel important for the virus lifecycle. The proton-channel activity is inhibited by amine-containing adamantyl drugs amantadine and rimantadine, which have been shown to bind specifically to the pore of M2TM near Ser31. However, whether the polar amine points to the N- or C-terminus of the channel has not yet been determined.

Conformational plasticity of the influenza A M2 transmembrane helix in lipid bilayers under varying pH, drug binding, and membrane thickness.

Membrane proteins change their conformations to respond to environmental cues, thus conformational plasticity is important for function. The influenza A M2 protein forms an acid-activated proton channel important for the virus lifecycle. Here we have used solid-state NMR spectroscopy to examine the conformational plasticity of membrane-bound transmembrane domain of M2 (M2TM).

Structure of the amantadine binding site of influenza M2 proton channels in lipid bilayers.

The M2 protein of influenza A virus is a membrane-spanning tetrameric proton channel targeted by the antiviral drugs amantadine and rimantadine. Resistance to these drugs has compromised their effectiveness against many influenza strains, including pandemic H1N1. A recent crystal structure of M2(22-46) showed electron densities attributed to a single amantadine in the amino-terminal half of the pore, indicating a physical occlusion mechanism for inhibition.

Effects of amantadine on the dynamics of membrane-bound influenza A M2 transmembrane peptide studied by NMR relaxation.

The molecular motions of membrane proteins in liquid-crystalline lipid bilayers lie at the interface between motions in isotropic liquids and in solids. Specifically, membrane proteins can undergo whole-body uniaxial diffusion on the microsecond time scale. In this work, we investigate the (1)H rotating-frame spin-lattice relaxation (T (1rho)) caused by the uniaxial diffusion of the influenza A M2 transmembrane peptide (M2TMP), which forms a tetrameric proton channel in lipid bilayers.

Structure and function of the influenza A M2 proton channel.

The M2 protein of influenza A viruses forms a tetrameric pH-activated proton-selective channel that is targeted by the amantadine class of antiviral drugs. Its ion channel function has been extensively studied by electrophysiology and mutagenesis; however, the molecular mechanism of proton transport is still elusive, and the mechanism of inhibition by amantadine is controversial. We review the functional data on proton channel activity, molecular dynamics simulations of the proton conduction mechanism, and high-resolution structural and dynamical information of this membrane protein in lipid bilayers and lipid-mimetic detergents.

Immobilization of the influenza A M2 transmembrane peptide in virus envelope-mimetic lipid membranes: a solid-state NMR investigation.

The dynamic and structural properties of membrane proteins are intimately affected by the lipid bilayer. One property of membrane proteins is uniaxial rotational diffusion, which depends on the membrane viscosity and thickness. This rotational diffusion is readily manifested in solid-state NMR spectra as characteristic line shapes and temperature-dependent line narrowing or broadening.

Discovery of spiro-piperidine inhibitors and their modulation of the dynamics of the M2 proton channel from influenza A virus.

Amantadine has been used for decades as an inhibitor of the influenza A virus M2 protein (AM2) in the prophylaxis and treatment of influenza A infections, but its clinical use has been limited by its central nervous system (CNS) side effects as well as emerging drug-resistant strains of the virus. With the goal of searching for new classes of M2 inhibitors, a structure-activity relation study based on 2-[3-azaspiro(5,5)undecanol]-2-imidazoline (BL-1743) was initiated. The first generation BL-1743 series of compounds has been synthesized and tested by two-electrode voltage-clamp (TEV) assays.

Accurate measurement of methyl 13C chemical shifts by solid-state NMR for the determination of protein side chain conformation: the influenza a M2 transmembrane peptide as an example.

The use of side chain methyl (13)C chemical shifts for the determination of the rotameric conformation of Val and Leu residues in proteins by solid-state NMR spectroscopy is described. Examination of the solution NMR stereospecifically assigned methyl groups shows significant correlation between the difference in the two methyl carbons' chemical shifts and the side chain conformation. It is found that alpha-helical and beta-sheet backbones cause different side chain methyl chemical shift trends.

Structure of amantadine-bound M2 transmembrane peptide of influenza A in lipid bilayers from magic-angle-spinning solid-state NMR: the role of Ser31 in amantadine binding.

The M2 proton channel of influenza A is the target of the antiviral drugs amantadine and rimantadine, whose effectiveness has been abolished by a single-site mutation of Ser31 to Asn in the transmembrane domain of the protein. Recent high-resolution structures of the M2 transmembrane domain obtained from detergent-solubilized protein in solution and crystal environments gave conflicting drug binding sites. We present magic-angle-spinning solid-state NMR results of Ser31 and a number of other residues in the M2 transmembrane peptide (M2TMP) bound to lipid bilayers.

Amantadine-induced conformational and dynamical changes of the influenza M2 transmembrane proton channel.

The M2 protein of influenza A virus forms a transmembrane proton channel important for viral infection and replication. Amantadine blocks this channel, thus inhibiting viral replication. Elucidating the high-resolution structure of the M2 protein and its change upon amantadine binding is crucial for designing antiviral drugs to combat the growing resistance of influenza A viruses against amantadine.

Simultaneous extraction of multiple orientational constraints of membrane proteins by 13C-detected N-H dipolar couplings under magic angle spinning.

A 13C-detected N-H dipolar coupling technique is introduced for uniaxially mobile membrane proteins for orientation determination using unoriented samples. For proteins undergoing rigid-body uniaxial rotation in the lipid bilayer, the intrinsic equality between the dipolar coupling constants measured in unoriented samples and the anisotropic coupling measured in static oriented samples has been shown recently. Here, we demonstrate that the orientation-sensitive backbone N-H dipolar couplings can be measured with 13C detection using 2D and 3D MAS correlation experiments, so that maximal site resolution can be achieved and multiple orientational constraints can be extracted from each experiment.

Determining the orientation of uniaxially rotating membrane proteins using unoriented samples: a 2H, 13C, AND 15N solid-state NMR investigation of the dynamics and orientation of a transmembrane helical bundle.

Membrane protein orientation has traditionally been determined by NMR using mechanically or magnetically aligned samples. Here we show a new NMR approach that abolishes the need for preparing macroscopically aligned membranes. When the protein undergoes fast uniaxial rotation around the bilayer normal, the 0 degrees -frequency of the motionally averaged powder spectrum is identical to the frequency of the aligned protein whose alignment axis is along the magnetic field.