Plasmid-encoded ropiness production inLactobacillus casei SSP.casei

Summary Genetic determinants of the Muc⁺ character were investigated in two ropy strains,Lactobacillus delbrueckii ssp.bulgaricus 201 andL. casei ssp.casei NCIB 4114, which secrete a large amount of slime in culture media. Plasmid DNA analysis revealed the presence of two plasmids (4.5 and 2.3 Mdal) inL. casei ssp.casei, whileL. delbrueckii ssp.bulgaricus was plasmid free, suggesting a chromosomal location of Muc⁺ character in this strain. Curing experiments carried out onL. casei ssp.casei NCIB 4114 indicated a correlation between the Muc⁺ phenotype and the 4.5 Mdal plasmid.     

Cytometry and flow cytometry of 4',6-diamidino-2-phenylindole (DAPI)-stained suspensions of nuclei released from fresh and fixed tissues of plants

Cytometry and flow cytometry were used to study characteristics of fluorescence of the DNA-DAPI complex in nuclei released from different fresh and formaldehyde-fixed pea ( Pisum sativum L. cv. Lincoln) tissues. The two methods of isolation are compared and discussed as well as their possible use for quantitative analysis of DNA in plant tissues. With fixed tissues it is possible to obtain a number of nuclei sufficient for the flow cytometric analysis, even using small amounts of plant tissue.     

Factors regulating baculovirus late and very late gene expression in transient-expression assays.

Eighteen genes of Autographa californica nuclear polyhedrosis virus are necessary and sufficient to transactivate expression from the late vp39 promoter in transient-expression assays in SF-21 cells. These 18 genes, known as late expression factor genes (lefs), are also required to transactivate the very late promoter of the polyhedrin gene, polh, but expression from this promoter is relatively weak compared with expression from the vp39 promoter. To further define the factors required for late and very late promoter expression, we first determined that the eighteen lefs were also required for expression from two other major baculovirus promoters: the late basic 6.9-kDa protein gene, p6.9, and the very late 10-kDa protein gene, p10. We next examined the effect of the very late expression factor 1 gene (vlf-1), a gene previously identified by analysis of a temperature-sensitive mutant, in the transient expression assay and found that vlf-1 specifically transactivated the two very late promoters but not the two late promoters. We then surveyed the Autographa californica nuclear polyhedrosis virus genome for additional genes which might specifically regulate very late gene expression; no additional vlf genes were detected, suggesting that VLF-1 is the primary regulator of very late gene expression. Finally, we found that the relative contribution of the antiapoptosis gene p35, which behaves as a lef in these transient-expression assays, depended on the nature of the other viral genes provided in the cotransfection mixtures, suggesting that other viral genes also contribute to the ability of the virus to block apoptosis.     

15N natural abundance in forest and pasture soils of the Brazilian Amazon Basin

The natural abundance of ¹⁵N was examined in soil profiles from forests and pastures of the Brazilian Amazon Basin to compare tropical forests on a variety of soil types and to investigate changes in the sources of nitrogen to soils following deforestation for cattle ranching. Six sites in the state of Rondônia, two sites in Pará and one in Amazonas were studied. All sites except one were chronosequences and contained native forest and one or more pastures ranging from 2 to 27 years old. Forest soil ¹⁵N values to a depth of 1 m ranged from 8 to 23 and were higher than values typically found in temperate forests. A general pattern of increasing ¹⁵N values with depth near the soil surface was broadly similar to patterns in other forests but a decrease in ¹⁵N values in many forest profiles between 20 and 40 cm suggests that illuviation of ¹⁵N-depleted nitrate may influence total soil ¹⁵N values in deeper soil where total N concentrations are low. In four chronosequences in Rondônia, the ¹⁵N values of surface soil from pastures were lower than in the original forest and ¹⁵N values were increasingly depleted in older pastures. Inputs of atmospheric N by dinitrogen fixation could be an important N source in these pastures. Other pastures in Amazonas and Pará and Rondônia showed no consistent change from forest values. The extent of fractionation that leads to ¹⁵N enrichment in soils was broadly similar over a wide range of soil textures and indicated that similar processes control N fractionation and loss under tropical forest over a broad geographic region. Forest ¹⁵N profiles were consistent with conceptual models that explain enrichment of soil ¹⁵N values by selective loss of ¹⁴N during nitrification and denitrification.     

In vivo and in vitro analyses of recombinant baculoviruses lacking a functional cg30 gene.

The cg30 gene of Autographa californica nuclear polyhedrosis virus (AcMNPV) encodes two sequence motifs, a zinc finger-like motif and a leucine zipper, found in other polypeptides known to be involved in gene regulation. To gain insight into the function of the cg30 product, CG30, we constructed and characterized recombinant viruses lacking a functional cg30 gene. We found that cg30 mutants had no striking phenotype in cell lines derived from Spodoptera frugiperda or Trichoplusia ni or in T. ni larvae. Although cg30 is known to be transcribed as an early monocistronic RNA and as the second cistron of an abundant late bicistronic RNA, production of a CG30-beta-galactosidase fusion protein was observed mainly at early times postinfection. Viruses containing cg30 had a subtle growth advantage over those lacking cg30 after several viral passages in cell culture. We employed transient expression assays to determine whether cg30 and pe-38, an AcMNPV gene that encodes a polypeptide with zinc finger-like and leucine zipper motifs similar to those of cg30, have redundant functions. Although pe-38 may have a role in AcMNPV gene expression, there was no indication that cg30 and pe-38 are functionally redundant.     

A baculovirus gene involved in late gene expression predicts a large polypeptide with a conserved motif of RNA polymerases.

We have identified and sequenced a novel baculovirus gene, late expression factor eight gene (lef-8), of Autographa californica nuclear polyhedrosis virus that is necessary for efficient expression from late and very late virus gene promoters in a transient expression assay. The predicted gene product, LEF-8, has a molecular mass of 102 kDa and contains a conserved sequence motif, GXKX4HGQ/NKG, found in DNA-directed RNA polymerases throughout the animal, plant, and microbial kingdoms.     

Eighteen baculovirus genes, including lef-11, p35, 39K, and p47, support late gene expression.

We report the identification of four additional genes of the Autographa californica nuclear polyhedrosis virus involved in expression from a late baculovirus promoter in transient expression assays. Three of these genes, p35, 39K, and p47, have been previously described. The role of the p35 gene product in late gene expression may be related to its ability to block apoptosis, since two other baculovirus genes also known to block apoptosis, Cp-iap and Op-iap, were able to functionally replace p35 in the transient expression assay. The requirement for p47 in this assay confirms its role in late gene expression, a role previously established by characterization of a temperature-sensitive mutant of p47, while the requirement for 39K may be related to its known association with the virogenic stroma. The fourth gene identified as a late expression factor gene, lef-11, was located immediately upstream of 39K and is predicted to encode a 13-kDa polypeptide. When plasmids containing these 4 genes were cotransfected with plasmids containing the 14 genes previously identified as late gene expression factors, the level of expression from the late capsid promoter was similar to that observed for a library of clones representing the entire viral genome. The genes provided by these 18 plasmids thus represent the viral genes necessary and sufficient to support expression from a late viral promoter in this transient expression assay.     

A chlorinated phenylpyrrole antibiotic fromActinoplanes

Two chlorophenyl-containing antibiotics have been isolated from a strain ofActinoplanes (ATCC 33002). Antibiotic A 15104 Y is a chlorinated phenylpyrrole compound whose structure has been confirmed by chemical synthesis. Antibiotic A 15104 Z is a chlorophenol derivative for which a structural formula is proposed on the basis of its physicochemical properties. A 15104 Y shows antimicrobial activity against Gram-positive, Gram-negative, and acid-fast bacteria, yeasts, fungi, and protozoa, while A 15104 Z possesses a low activity against Gram-positive bacteria andTrichophyton. A 15104 Y has a weak activity in curing experimentally infected mice, at a dose that is one-fifth the LD₅₀ for the same species. This is the first example of a chloropyrrole derivative isolated from an actinomycete.     

Inhibition of elongation and H+ and K+ transport by the phosphodiesterase inhibitor aminophylline in plant tissue

Aminophylline, an inhibitor of cyclic nucleotide phosphodiesterase (EC 3.1.4.17), inhibits elongation and correlated H⁺ and K⁺ transport in embryos of Haplopappus gracilis and in pea internode segments. Moreover, the drug strongly inhibits the stimulation of these processes by fusicoccin and indole-3-acetic acid and reduces passive permeability of the membrane. The possible mechanisms of action of aminophylline are discussed.Abbreviations cAMP adenosine 3:5-cyclic monophosphate - FC fusicoccin - IAA indole-3-acetic acid - MES 2-N-morpholinoethanesulfonic acid - PDE cyclic nucleotide phosphodiesterase