Mutagen sensitivity and suppression of position-effect variegation result from mutations in mus209, the Drosophila gene encoding PCNA.

The mus209B1 mutant of Drosophila melanogaster exhibits a complex pleiotropy of temperature-sensitive (ts) lethality, hypersensitivity to DNA-damaging agents such as ionizing radiation and methyl methanesulfonate, suppression of position-effect variegation (PEV), and female sterility. Our discovery that mus209 encodes proliferating cell nuclear antigen (PCNA), which is an indispensable component of the DNA replication apparatus, suggests that alterations to chromosome replication may underlie that pleiotropy. Nine lethal mutations, three of them ts, genetically define the Pcna locus. Temperature shift studies reveal that the vital function of PCNA is required throughout virtually all stages of fly development, and that maternally encoded PCNA is essential for embryogenesis. All three ts mutants strongly suppress PEV, which suggests a role for PCNA in chromatin assembly or modification.     

Identity of the RNA-binding protein K of hnRNP particles with protein H16, a sequence-specific single strand DNA-binding protein.

Protein H16, which we have identified previously in mammalian cell lines, binds in vitro to two single stranded DNA sites on the late strand of the early promoter of SV40. It has no other single strand binding site in the SV40 genome and does not bind to double stranded DNA. In vitro, H16 can be shown to stimulate strongly the activity of purified RNA polymerase II. Here we have purified this 70 kDa protein from cultured monkey cells and have sequenced three of its tryptic peptides. The analysis indicates that H16 is the simian homolog of human protein K, a nuclear RNA-binding protein found in heterogeneous nuclear ribonucleoprotein (hnRNP) particles, which contains a KH domain present in several proteins including the fragile X mental retardation gene product (FMR1). The binding affinities of protein K/H16 for RNA and DNA were subsequently compared in detail. They showed that under conditions where K/H16 binds strongly to its single stranded DNA site, it binds very weakly to the corresponding RNA sequence. This result suggests a possible shuttling of the protein from RNA to DNA during processes which involve opening of the DNA double helix.     

The alphaviruses: gene expression, replication, and evolution.

The alphaviruses are a genus of 26 enveloped viruses that cause disease in humans and domestic animals. Mosquitoes or other hematophagous arthropods serve as vectors for these viruses. The complete sequences of the +/- 11.7-kb plus-strand RNA genomes of eight alphaviruses have been determined, and partial sequences are known for several others; this has made possible evolutionary comparisons between different alphaviruses as well as comparisons of this group of viruses with other animal and plant viruses. Full-length cDNA clones from which infectious RNA can be recovered have been constructed for four alphaviruses; these clones have facilitated many molecular genetic studies as well as the development of these viruses as expression vectors. From these and studies involving biochemical approaches, many details of the replication cycle of the alphaviruses are known. The interactions of the viruses with host cells and host organisms have been exclusively studied, and the molecular basis of virulence and recovery from viral infection have been addressed in a large number of recent papers. The structure of the viruses has been determined to about 2.5 nm, making them the best-characterized enveloped virus to date. Because of the wealth of data that has appeared, these viruses represent a well-characterized system that tell us much about the evolution of RNA viruses, their replication, and their interactions with their hosts. This review summarizes our current knowledge of this group of viruses.     

Nucleocapsid-glycoprotein interactions required for assembly of alphaviruses.

We have studied interactions between nucleocapsids and glycoproteins required for budding of alphaviruses, using Ross River virus-Sindbis virus chimeras in which the nucleocapsid protein is derived from one virus and the envelope glycoproteins are derived from the second virus. A virus containing the Ross River virus genome in which the capsid protein had been replaced with that from Sindbis virus was almost nonviable. Nucleocapsids formed in normal numbers in the infected cell, but very little virus was released from the cell. There are 11 amino acid differences between Ross River virus and Sindbis virus in their 33-residue E2 cytoplasmic domains. Site-specific mutagenesis was used to change 9 of these 11 amino acids in the chimera from the Ross River virus to the Sindbis virus sequence in an attempt to adapt the E2 of the chimera to the nucleocapsid. The resulting mutant chimera grew 4 orders of magnitude better than the parental chimeric virus. This finding provides direct evidence for a sequence-specific interaction between the nucleocapsid and the E2 cytoplasmic domain during virus budding. The mutated chimeric virus readily gave rise to large-plaque variants that grew almost as well as Ross River virus, suggesting that additional single amino acid substitutions in the structural proteins can further enhance the interactions between the disparate capsid and the glycoproteins. Unexpectedly, change of E2 residue 394 from lysine (Ross River virus) to glutamic acid (Sindbis virus) was deleterious for the chimera, suggesting that in addition to its role in nucleocapsid-E2 interactions, the N-terminal part of the E2 cytoplasmic domain may be involved in glycoprotein-glycoprotein interactions required to assemble the glycoprotein spikes. The reciprocal chimera, Sindbis virus containing the Ross River virus capsid, also grew poorly. Suppressor mutations arose readily in this chimera, producing a virus that grew moderately well and that formed larger plaques.     

The ecological significance of nickel hyperaccumulation: a plant chemical defense

Nickel hyperaccumulating plants have more than 1000 mg Ni kg^–1 dry weight when grown on nickel-bearing soils. We hypothesized that Ni hyperaccumulation could serve as a chemical defense against herbivores In feeding experiments with potential insect herbivores and Ni hyperaccumulating plants, only those inseets fed leaves from plants grown on non-nickel-bearing soil survived or showed a weight gain. Among chemical parameters measured, only Ni content of plants was sufficient to explain this result. When subjected to herbivory by lepidopteran larvae, plants grown on Ni-amended soil showed greater survival and yield than plants on unamended soil. Ni hyperaccumulation may be an effective plant chemical defense against herbivores because of its high lethality, apparent low cost, and broad spectrum of toxicity.     

Assignment of the gene encoding the human thyrotropin-releasing hormone receptor to 8q23 by fluorescence in situ hybridization

A cDNA for human thyrotropin-releasing hormone (TRH) receptor has been isolated from a human pituitary cDNA library. By using this cDNA as a biotinylated probe, the gene encoding the TRH receptor has been localized to chromosome 8q23 by in situ hybridization.     

The spatial variability of soil resources following long-term disturbance

The spatial distributions of selected soil properties in two adjacent sites in southwest Michigan were examined to evaluate the potential effects of chronic disturbance on resource heterogeneity. One site was a cultivated field that had been cleared, plowed, and cropped annually for decades prior to sampling while the other, uncultivated field was cleared of original forest in 1960 after which it was mown annually but never plowed or cropped. We took replicate samples from a 330-point unaligned grid across the sites for soil pH, gravimetric moisture, inorganic phosphorus, total carbon, and net nitrification and nitrogen mineralization potentials. Soils in the cultivated site contained less than half as much carbon as in the uncultivated site, but had higher levels of inorganic phosphorus and moisture, and higher soil pH. Potential net nitrogen mineralization and nitrification rates did not differ between sites. Geostatistical analysis showed that almost all properties examined were strongly autocorelated within each site; structural variance as a proportion of sample variance ranged from 30–95% for all properties, and for any given property differed little between sites. The distance over which this dependence was expressed, however, was for all properties but pH substantially less in the uncultivated site (7–26 m) as compared to the tilled site (48–108m), especially for total C and net nitrification and N mineralization. These results suggest that the spatial pattern and scale of soil variability can differ markedly among edaphically identical sites and that these differences can be related to disturbance history.     

ANTIVIRAL ACTIVITY OF CULTURED BLUE-GREEN ALGAE (CYANOPHYTA)1

Lipophilic and hydrophilic extracts from approximately 600 strains of cultured cyanophytes, representing some 300 species, were examined for antiviral activity against three pathogenic viruses. Approximately 10% of the cultures produced substances that caused significant reduction in cytopathic effect normally associated with viral infection. The screening program identified the order Chroococcales as commonly producing antiviral agents.