Chromosome segregation: playing polo in prophase

During mitosis, in most eukaryotes, cohesin is removed from chromosomes in two steps. A paper in the March issue of Molecular Cell identifies Polo-like kinase as a key regulator for the first step that releases much of cohesin during prophase.     

DelGEF, an RCC1-related protein encoded by a gene on chromosome 11p14 critical for two forms of hereditary deafness

We have cloned a human cDNA, DELGEF (deafness locus associated putative guanine nucleotide exchange factor), derived from a 225 kb genomic sequence of chromosome 11p14, critical for the Usher 1C syndrome and for DFNB18, a locus for non-syndromic sensorineural deafness. The amino acid sequence of the protein hDelGEF1 is homologous to the nucleotide exchange factor RCCI for the small GTPase Ran. hDelGEF2 is derived from the same DELGEF gene by alternative splicing. In addition, we have identified a murine homologue, mDelGEF. The ubiquitously expressed soluble protein hDelGEF1 is found both in the cyytoplasm and in the nucleus. Overexpressed hDelGEF2 colocalizes with mitochondria.     

Increasement of Heterologous Expression of Recombinant Vit v 1 in Pichia pastoris KM71 by Nonionic Detergents as a Cost-effective Approach

Applied Biochemistry and Microbiology - Vit v 1 as a lipid-transfer protein is a major allergen of grapes (Vitis vinifera) that elicits food allergy in many patients in Iran. Todays, recombinant...     

Conversion of Unprotected Amino-Link Oligonucleotides into Their Tetramethylguanidinium Derivatives

Abstract

The preparation of tetramethylguanidinium oligodeoxynucleotide (ODN) derivatives by reaction of the corresponding aminoalkyl-ODN with the uronium salts HBTU, TBTU or HATU, respectively, is described. The binding affinity of the new tetramethylguanidinium ODN derivatives was determined.     

Synthesis of 3′,5′-Dithymidylyl-α-hydroxyphosphonate Dimer Building Blocks for Oligonucleotide Synthesis—A New Pro-oliguncleotide

Abstract

The synthesis of the dimer building blocks 1 and 2 and their introduction into (T)₁₅-oligonucleotides is described. The stability against 3′-exonuclease digestion (SVP) as well as the hybridization properties (T_m values) were examined.     

Epidemiology of No-Code Orders in an Academic Hospital

Relatively little is known about the circumstances in which decisions not to resuscitate, documented by no-code orders, are made. By review of medical records and interviews with house staff officers, we studied all medical service patients for whom no-code orders were written and those patients who received cardiopulmonary resuscitation (CPR) between October and December 1980 in the Portland Veterans Administration Medical Center.Among 1,780 patients admitted, 56 (3.1%) received no-code orders. All decisions were reportedly made by groups of individuals usually including the intern (98% of cases) and resident (93%), but not attending physician (39%). Many patients (43%) were disoriented or obtunded at the time of the no-code decision and 80% of oriented patients did participate in the decision.Thirty-seven of the 56 no-code patients died during the study. Comparing these with 20 patients who experienced cardiac arrest and did receive CPR, cancer, dementia, incontinence, non-ambulatory, divorced-separated and unemployed statuses were all more prevalent among no-code patients (P<.05).No-code orders in this Veterans Administration teaching hospital were relatively common and appeared to be made collectively. Participation of patients and attending physicians in the decisions, however, was limited.     

Mimicking a cytoskeleton by coupling poly(N-isopropylacrylamide) to the inner leaflet of liposomal membranes: effects of photopolymerization on vesicle shape and polymer architecture

Networks of N-isopropylacrylamide (NIPAM) copolymers, coupled to spherical phospholipid bilayers, are suitable as a model for the study of the interaction between the cytoskeleton and cellular membranes, as well as for promising new drug delivery systems with triggerable drug release properties and improved stability. In this article, we describe a simple preparation technique for liposomes from egg phosphatidyl choline (EPC) encapsulating a cross-linked NIPAMminus signTEGDM copolymer skeleton (tetraethylene glycol dimethacrylate, TEGDM) which is coupled only to the inner monolayer by a novel membrane anchor monomer. Polymerization in the lipid vesicles was initiated at the inner membrane surface by the radical initiator 2,2-diethoxy-acetophenone (DEAP) permeating through the membrane from the outside. The effects of photopolymerization and polymer formation on vesicle shape and membrane integrity were studied by transmission electron microscopy (TEM), cryo-TEM, and atomic force microscopy (AFM). Upon UV irradiation, approximately 100% of the vesicles contained a polymer gel and only occasional changes in the spherical shape of the liposomes were observed. The architecture of the polymer network inside the liposomal compartment was determined by the conditions of the photopolymerization. Composite structures of polymer hollow spheres or solid spheres, respectively, tethered to spherical membrane vesicles were produced. The increased stability of the polymer-tethered lipid bilayers against solubilization by sodium cholate, compared to pure EPC vesicles, was determined by radiolabeling the lipid membrane.     

Identification and quantification of GC-rich oligodeoxynucleotides in tissue extracts by capillary gel electrophoresis

Capillary gel electrophoresis (CGE) is a widely used method for quantification of oligonucleotide-based drugs, such as CpG oligodeoxynucleotides (CpG ODN), aptamers and small interfering ribonucleic acids (siRNAs) that allows accurate quantification of parent compound as well as metabolites. Stable secondary structure formation of these molecules frequently prevents analysis by conventional CGE methods and impedes pharmacokinetic assessment. Herein, we describe development of a CGE method for identification and quantification of complex mixtures of secondary structure forming GC-rich ODN in biological samples at dose levels of 0.5mg/kg and above. Samples containing GC-rich CpG ODN and metabolite markers were treated by solid-phase-extraction (SPE) and subsequently analyzed by CGE using a 50cm neutrally coated capillary at 60 degrees C together with a 7M urea buffer system containing 30% dimethylsulfoxide (DMSO). Peak resolutions >or=1 were typically achieved, enabling pharmacokinetic assessment of secondary structure forming oligonucleotides in biological samples that hitherto were unsusceptible to quantitative analysis.     

Cell-to-cell and phloem-mediated transport of potato virus X. The role of virions

Movement-deficient potato virus X (PVX) mutants tagged with the green fluorescent protein were used to investigate the role of the coat protein (CP) and triple gene block (TGB) proteins in virus movement. Mutants lacking either a functional CP or TGB were restricted to single epidermal cells. Microinjection of dextran probes into cells infected with the mutants showed that an increase in the plasmodesmal size exclusion limit was dependent on one or more of the TGB proteins and was independent of CP. Fluorescently labeled CP that was injected into epidermal cells was confined to the injected cells, showing that the CP lacks an intrinsic transport function. In additional experiments, transgenic plants expressing the PVX CP were used as rootstocks and grafted with nontransformed scions. Inoculation of the PVX CP mutants to the transgenic rootstocks resulted in cell-to-cell and systemic movement within the transgenic tissue. Translocation of the CP mutants into sink leaves of the nontransgenic scions was also observed, but infection was restricted to cells close to major veins. These results indicate that the PVX CP is transported through the phloem, unloads into the vascular tissue, and subsequently is transported between cells during the course of infection. Evidence is presented that PVX uses a novel strategy for cell-to-cell movement involving the transport of filamentous virions through plasmodesmata.