Cancellation of cellular responses to nanoelectroporation by reversing the stimulus polarity

Nanoelectroporation of biomembranes is an effect of high-voltage, nanosecond-duration electric pulses (nsEP). It occurs both in the plasma membrane and inside the cell, and nanoporated membranes are distinguished by ion-selective and potential-sensitive permeability. Here we report a novel phenomenon of bioeffects cancellation that puts nsEP cardinally apart from the conventional electroporation and electrostimulation by milli- and microsecond pulses. We compared the effects of 60- and 300-ns monopolar, nearly rectangular nsEP on intracellular Ca²⁺ mobilization and cell survival with those of bipolar 60 + 60 and 300 + 300 ns pulses. For diverse endpoints, exposure conditions, pulse numbers (1–60), and amplitudes (15–60 kV/cm), the addition of the second phase cancelled the effects of the first phase. The overall effect of bipolar pulses was profoundly reduced, despite delivering twofold more energy. Cancellation also took place when two phases were separated into two independent nsEP of opposite polarities; it gradually tapered out as the interval between two nsEP increased, but was still present even at a 10-µs interval. The phenomenon of cancellation is unique for nsEP and has not been predicted by the equivalent circuit, transport lattice, and molecular dynamics models of electroporation. The existing paradigms of membrane permeabilization by nsEP will need to be modified. Here we discuss the possible involvement of the assisted membrane discharge, two-step oxidation of membrane phospholipids, and reverse transmembrane ion transport mechanisms. Cancellation impacts nsEP applications in cancer therapy, electrostimulation, and biotechnology, and provides new insights into effects of more complex waveforms, including pulsed electromagnetic emissions.     

Helminthosporoside,a host-specific toxin fromHelminthosporium sacchari: A structure revision and a new partial structure

Summary Reevaluation of the previously proposed structure of helminthosporoside, a host-specific toxin fromHelminthosporium sacchari, reveals a sesquiterpenoid bis-digalactoside. The carbohydrate portion of the toxin was characterized by¹³C-NMR spectroscopy, methylation analyses, FD and FAB mass spectroscopy. The ring size and anomeric configuration of the galactose moieties were determined by utilizing a¹³C-NMR structural analysis method. A new partial structure is proposed.Acknowledgment. We extend special thanks to C.E. Costello, Department of Chemistry, M.I.T., Cambridge, MA, for FD mass spectral analyses, and to C.E. Ballou, Department of Biochemistry, University of California, Berkeley, CA, for FAB mass spectral analyses. We submit sincere appreciation to both V.N. Reihold, Harvard Medical School, Boston, MA, and P. Albersheim, Department of Chemistry, University of Colorado, Boulder, CO, for methylation analyses. We thank J.S. Frye at the Colorado State University Regional NMR-Center, funded by National Science Foundation grant No. DHE 78-18581, Department of Chemistry, Colorado State University, Fort Collins, Co, for high resolution CMR and NMR, and we are grateful to P.W. Jennings, Department of Chemistry, Montana State University, Bozeman, Mt, for total carbon analyses. This work was supported in part by a Herman Frasch Foundation grant to G.A. Strobel, by NSF grant PCM-78-22517, and funds from the Montana Agricultural Experiment Station. B.P. Mundy acknowledges the partial support of NSF (ISP-801149).     

大学体育口号与理念的谱系与解构——以清华大学为例

体育口号与理念作为体育思想的一种表现形式,是学校开展体育工作的手段之一。研究选取大学体育口号与理念为研究对象,通过文献资料法、问卷调查法和专家访谈法,梳理大学体育口号与理念的历史谱系,以不同时期清华大学的体育口号与理念为个案分析,审视我国大学体育思想的发展脉络。研究指出,从“为祖国健康工作五十年”到“无体育,不清华”体育口号与理念的谱系,体现了从“卡里斯马”到“集体无意识”的价值导入,呈现出大学体育价值理念的文化记忆。大学体育口号与理念的内涵表现在：从中华民族的救国到建国再到强国的爱国性,从体育运动的强迫到认同再到自发的传承性,从影响范围的区域到国家再到国际的卓越性以及对大学体育的引领性。研究认为,大学体育口号与理念体现了大学体育价值取向的变化以及大学体育思想的继承发扬,是实现“体育的迁移价值”和立德树人的有效手段。     

THM1 negatively modulates mouse sonic hedgehog signal transduction and affects retrograde intraflagellar transport in cilia

Characterization of previously described intraflagellar transport (IFT) mouse mutants has led to the proposition that normal primary cilia are required for mammalian cells to respond to the sonic hedgehog (SHH) signal. Here we describe an N-ethyl-N-nitrosourea-induced mutant mouse, alien (aln), which has abnormal primary cilia and shows overactivation of the SHH pathway. The aln locus encodes a novel protein, THM1 (tetratricopeptide repeat-containing hedgehog modulator-1), which localizes to cilia. aln-mutant cilia have bulb-like structures at their tips in which IFT proteins (such as IFT88) are sequestered, characteristic of Chlamydomonas reinhardtii and Caenorhabditis elegans retrograde IFT mutants. RNA-interference knockdown of Ttc21b (which we call Thm1 and which encodes THM1) in mouse inner medullary collecting duct cells expressing an IFT88-enhanced yellow fluorescent protein fusion recapitulated the aln-mutant cilial phenotype, and live imaging of these cells revealed impaired retrograde IFT. In contrast to previously described IFT mutants, Smoothened and full-length glioblastoma (GLI) proteins localize to aln-mutant cilia. We hypothesize that the aln retrograde IFT defect causes sequestration of IFT proteins in aln-mutant cilia and leads to the overactivated SHH signaling phenotype. Specifically, the aln mutation uncouples the roles of anterograde and retrograde transport in SHH signaling, suggesting that anterograde IFT is required for GLI activation and that retrograde IFT modulates this event.     

Efficient generation and mapping of recessive developmental mutations using ENU mutagenesis

Treatment with N-ethyl-N-nitrosourea (ENU) efficiently generates single-nucleotide mutations in mice. Along with the renewed interest in this approach, much attention has been given recently to large screens with broad aims; however, more finely focused studies have proven very productive as well. Here we show how mutagenesis together with genetic mapping can facilitate the rapid characterization of recessive loci required for normal embryonic development. We screened third-generation progeny of mutagenized mice at embryonic day (E) 18.5 for abnormalities of organogenesis. We ascertained 15 monogenic mutations in the 54 families that were comprehensively analyzed. We carried out the experiment as an outcross, which facilitated the genetic mapping of the mutations by haplotype analysis. We mapped seven of the mutations and identified the affected locus in two lines. Using a hierarchical approach, it is possible to maximize the efficiency of this analysis so that it can be carried out easily with modest infrastructure and resources.     

ENVIRONMENTAL CHALLENGES FACING INDIA

INTRODUCTION India is among one of the ten most industrialized nations in the world. Increase in population has raised the urbar industrial, transport and agriculture demands which are major reasons for the degradation of the environmel condition. For India besides land and soil degradation, deforestation, low accessibility of water, ,industrial pollution and urban congestion are the major environmental issues of priority. The industries that generate huge quantities of waste are thermal power station, Iron and Steel Plants, Sugar, Paper and Fertilizer Industries.     

Deficiency of UBR1, a ubiquitin ligase of the N-end rule pathway, causes pancreatic dysfunction, malformations and mental retardation (Johanson-Blizzard syndrome)

Johanson-Blizzard syndrome (OMIM 243800) is an autosomal recessive disorder that includes congenital exocrine pancreatic insufficiency, multiple malformations such as nasal wing aplasia, and frequent mental retardation. We mapped the disease-associated locus to chromosome 15q14-21.1 and identified mutations, mostly truncating ones, in the gene UBR1 in 12 unrelated families with Johanson-Blizzard syndrome. UBR1 encodes one of at least four functionally overlapping E3 ubiquitin ligases of the N-end rule pathway, a conserved proteolytic system whose substrates include proteins with destabilizing N-terminal residues. Pancreas of individuals with Johanson-Blizzard syndrome did not express UBR1 and had intrauterine-onset destructive pancreatitis. In addition, we found that Ubr1(-/-) mice, whose previously reported phenotypes include reduced weight and behavioral abnormalities, had an exocrine pancreatic insufficiency, with impaired stimulus-secretion coupling and increased susceptibility to pancreatic injury. Our findings indicate that deficiency of UBR1 perturbs the pancreas' acinar cells and other organs, presumably owing to metabolic stabilization of specific substrates of the N-end rule pathway.