Determining the gluonic gravitational form factors of the proton.

The proton is one of the main building blocks of all visible matter in the Universe1. Among its intrinsic properties are its electric charge, mass and spin2. These properties emerge from the complex dynamics of its fundamental constituents-quarks and gluons-described by the theory of quantum chromodynamics3-5. The electric charge and spin of protons, which are shared among the quarks, have been investigated previously using electron scattering2. An example is the highly precise measurement of the electric charge radius of the proton6. By contrast, little is known about the inner mass density of the proton, which is dominated by the energy carried by gluons. Gluons are hard to access using electron scattering because they do not carry an electromagnetic charge. Here we investigated the gravitational density of gluons using a small colour dipole, through the threshold photoproduction of the J/ψ particle. We determined the gluonic gravitational form factors of the proton7,8 from our measurement. We used a variety of models9-11 and determined, in all cases, a mass radius that is notably smaller than the electric charge radius. In some, but not all cases, depending on the model, the determined radius agrees well with first-principle predictions from lattice quantum chromodynamics12. This work paves the way for a deeper understanding of the salient role of gluons in providing gravitational mass to visible matter.

Ultrafast photoinduced dynamics of halogenated cyclopentadienes: observation of geminate charge-transfer complexes in solution.

The photoinduced dynamics of the fully halogenated cyclopentadienes C5Cl6 and C5Br6 have been investigated in solution and gas phase by femtosecond time-resolved spectroscopy. Both in solution and in gas phase, homolytic dissociation into a halogen radical and a C5X5 (X = Cl, Br) radical was observed. In liquid phase, solvent-dependent formation of charge transfer complexes between geminate radicals was observed for the first time. These complexes were found to be surprisingly stable and offered the opportunity to follow the dynamics of specific radical pairs. In the case of C5Cl6 in trichloroethanol, a reaction of the chlorine radical with molecules from the solvent cage was observed.

mei-3, a recombination and repair gene of Neurospora crassa, encodes a RecA-like protein.

Neurospora crassa mei-3 is a mutant which exhibits meiotic and mitotic defects and mutagen sensitivity. Its defect is believed to be in recombination and repair. We have cloned the mei-3 gene from a N. crassa cosmid library of genomic DNA. Restriction fragment length polymorphism analysis determined the location of the cloned fragment was on chromosome one in approximately the same position that was previously reported for mei-3 by classical genetic methods. Deletion analysis showed the approximate coding region of mei-3 on the cloned genomic fragment. Northern blot analysis identified a 900-bp transcript. Sequencing revealed a 798-bp open reading frame with high coding preference which could encode a protein having a molecular weight of approximately 29,000. The predicted protein product of mei-3 has significant identity to the Rad51 and Dmc1 proteins from Saccharomyces cerevisiae. The mei-3 gene and both yeast genes have significant primary sequence homology with RecA, a recombination protein identified in Escherichia coli. The results suggest RecA-like proteins involved in DNA recombination and repair are highly conserved in eukaryotes.

The induction and repair of (6-4) photoproducts in Neurospora crassa.

The (6-4) photoproduct lesion found in DNA after UV irradiation is repaired by germinating Neurospora crassa conidia. Wild-type Neurospora removes 80% of the (6-4) photoproduct in approximately 20 min and maximal repair is accomplished by 30 min with approximately 89% of the original lesions removed. Mutagen-sensitive Neurospora mutants belonging to the established excision repair epistasis group, UVS-2, are not defective in the removal of cyclobutane pyrimidine dimers. Furthermore, we find these mutants capable of removing (6-4) photoproducts from their DNA at a rate similar to wild type. Comparable kinetics are also observed in key members of the other two epistasis groups.

The nucleotide excision repair epistasis group in Neurospora crassa.

DNA repair mutants in eucaryotes are normally assigned to three epistasis groups. Each epistasis group represents a "pathway" for DNA repair. The pathways are commonly designated (1) nucleotide excision repair, (2) recombination repair and (3) mutagenic repair. An excision repair epistasis group has been established in Neurospora and the mutants assigned to this group should be limited in their ability to excise pyrimidine dimers and other bulky lesions from DNA. Using a pyrimidine dimer-specific assay, we have found that all Neurospora crassa mutants assigned to the excision repair epistasis group are capable of removing pyrimidine dimers from the DNA at a rate similar to the wild-type organism.

Antiviral activities of gossypol and its derivatives against herpes simplex virus type II.

Gossypol, a disequiterpene obtained from cottonseed oil, and a series of peri-acylated gossylic nitriles were compared for their antiviral activities against HSV-II and for their toxicities to the host Vero cells. All of the peri-acylated gossylic nitriles exhibited lower cytotoxicities to the host cell than did the parent compound gossypol. Both gossypol and the series of derivatives exhibited antiviral activities against HSV-II when the virus was treated with drug at concentrations as low as 5 X 10(-7) M. Two of the derivatives, gossylic nitrile-1,1'-diacetate and gossylic nitrile-1,1'-divalerate, were capable of inhibiting viral multiplication in Vero cells that were infected with virus before administration of the drug. The results of this study indicate that modification of the aldehyde functional groups on gossypol lowers the toxicity of this drug but does not abolish its antiviral properties. Derivatives of gossypol may be useful antiviral agents.

The reversion of temperature-sensitive mutants of encephalomyocarditis virus upon serial passage in three continuous cell lines.

Several temperature-sensitive mutants of encephalomyocarditis virus were serially passed numerous times in HeLa, L929 and BHK-21 cells. The reversion of most of the mutants occurred most rapidly in BHK-21 cells and least rapidly in HeLa cells.

Defective interfering particles of encephalomyocarditis virus.

Encephalomyocarditis (EMC) virus and five temperature-sensitive mutants of EMC virus were serially passed numerous times in HeLa, L929 and BHK-21 cells. Equilibrium centrifugation in CsCl density gradients of virus and electrophoresis of virus RNA from many of the high number passages indicated the presence of standard EMC virus and particles containing RNA of lower molecular weight. Three virus isolates with high passage number were able to interfere with the replication of the standard virions in mixed infections. We believe our results show the generation of defective interfering particles of EMC virus.

Functional characterization of cleavage-defective mutants of encephalomyocarditis virus.

We characterized seven temperature-sensitive capsid cleavage (cleavage-defective) mutants of encephalomyocarditis virus. Our experimental approach was to monitor in vitro proteolysis reactions of either wild-type or cleavage-defective mutant capsid precursors mixed with cell-free translation products (containing the viral protease) of either wild-type or mutant viral RNA. The cell-free translation reactions and in vitro proteolysis reactions were done at 38 degrees C, because at this temperature cleavage of the capsid precursors was restricted in reactions containing cleavage-defective mutant viral RNA as the message, relative to those reactions containing wild-type viral RNA as the message. Wild-type or cleavage-defective mutant capsid precursors were prepared by adding cycloheximide to cell-free translation reactions primed with wild-type or mutant viral RNA, respectively, 12 min after the initiation of translation. In vitro proteolysis of wild-type capsid precursors with cell-free translation products of either wild-type or cleavage-defective mutant viral RNA led to similar products at 38 degrees C, indicating that the cleavage-defective mutant viral protease was not temperature sensitive. As a corollary to this, at 38 degrees C cleavage-defective mutant capsid precursors were not cleaved as completely as were wild-type capsid precursors by products of cell-free translation of wild-type viral RNA. The results from these in vitro proteolysis experiments indicate that all seven of the cleavage-defective mutants have capsid precursors with a temperature-sensitive configuration.

Two-dimensional electrophoretic analysis of encephalomyocarditis viral proteins.

All four capsid proteins of encephalomyocarditis virus and the precursor to two of these were resolved from purified virions with two-dimensional gel electrophoresis. In addition, all of the known stable virus-specific proteins found in infected cells, but not the primary and intermediate precursor proteins, could be resolved with these techniques.

Isolation and preliminary characterization of temperature-sensitive mutants of encephalomyocarditis virus.

Thirty temperature-sensitive mutants of encephalomyocarditis virus have been isolated and partially characterized. Fifteen of these mutants are phenotypically RNA+ thirteen are RNA-, and two are RNA +/-. Six RNA + mutants, one RNA- mutants, and one RNA +/- mutant have virions which are more thermosensitive at 56 degree C than the wild-type virions. Hela cells infected at the nonpermissive temperature with any of the RNA+ mutants produced neither infective nor noninfective viral particles. The cleavage of the precursor polypeptides in cells infected with 11 of the RNA+ mutants was defective at the nonpermissive temperature. This defect in cleavage occurred only in those precursor polypeptides leading to capsid proteins.

Studies of temperature sensitive mutants of bacteriophage Qbeta, defective in both replication and translation.

Temperature sensitive mutants of bacteriophage Qbeta have been isolated which fail in the synthesis of their virus RNA at the non-permissive temperature (42 degrees C). Nine mutants have been studied in some detail. Cells infected with these mutants at 37 degrees C and incubated long enough to produce substantial amounts of Qbeta RNA cease Qbeta RNA replication when shifted to 42 degrees C. The mutants can be classified into 3 groups according to the amount of Qbeta RNA replicase activity exhibited in extracts from infected cells isolated at various times after shift to 42 degrees C: in group 1 mutants, enzyme activity is the same, regardless of the time of isolation after shift; in group 2 mutants enzyme activity increases with time of isolation after shift; in group 3 mutants, enzyme activity decreases with time of isolation after shift. Synthesis of all virus proteins is suppressed at 42 degrees C in cells infected with group 2 of group 3 mutants. In cells infected with group 2 mutants, synthesis of Qbeta RNA replicase subunit beta is increased, but synthesis of other virus proteins is depressed at 42 degrees C. The inhibition of virus RNA and protein synthesis is reversible. A detailed analysis of these experiments suggests that a defective Qbeta RNA replicase is involved in the inhibition of both virus RNA and protein synthesis.

Electrophoretic and other properties of bacteriophage Q : the effect of a variable number of read-through proteins.

When subjected to electrophoresis in polyacrylamide gels, the virions of wild-type Qbeta bacteriophage are found in a single, major, anomalously wide band. With Qbeta mutant 27-2, this wide band is replaced by a set of narrow, well-defined bands. The most rapidly migrating band of the mutant, comprising less than 10% of the total, contains defective virions. These virions have sedimentation coefficients ranging from 70 to 100% of the bulk of the unfractionated mutant, they contain no read-through protein (protein IIb), and they are deficient in maturation protein and contain fragmented RNA. The second band, comprising less than 3% of the total virus, has not been well characterized. The virions in the remaining electrophoretic bands are infective. Their distribution into bands is believed due to differences in their effective volume resulting from differences in their content of protein IIb. The most rapidly migrating band of this series contains virions with a few molecules of IIb protein, whereas the more slowly migrating bands contain virions with a larger number of IIb molecules. The adjacent bands in the series contain virions which differ by approximately three IIb molecules. Wild-type Qbeta virus is similar to the mutant in that the more slowly migrating virions contain more protein IIb than the more rapidly migrating virions. Their failure to resolve into distinct bands upon electrophoresis is believed due to a less restricted packing of protein IIb into their virions. Both wild-type Qbeta and mutant 27-2 also have 1 to 5% of the virions in the form of dimers which migrate with approximately one-half the mobility of the respective monomer forms. When the average amount of IIb per virion is increased by growth of the virus in a UGA suppressor strain, the electrophoretic pattern is altered. In the case of wild-type Qbeta, the single band is wider, whereas with Qbeta mutant 27-2 there occurs an increased number of partially resolved narrow bands. We suggest that the structural feature responsible for the difference in electrophoretic pattern between mutant 27-2 and wild-type Qbeta is the mode of IIb packing in the virions. In the mutant, the IIb proteins are found in the virions only in multiples of three, whereas wild-type virions may differ by only a single IIb protein.