The genome of the social amoeba Dictyostelium discoideum.

The social amoebae are exceptional in their ability to alternate between unicellular and multicellular forms. Here we describe the genome of the best-studied member of this group, Dictyostelium discoideum. The gene-dense chromosomes of this organism encode approximately 12,500 predicted proteins, a high proportion of which have long, repetitive amino acid tracts. There are many genes for polyketide synthases and ABC transporters, suggesting an extensive secondary metabolism for producing and exporting small molecules. The genome is rich in complex repeats, one class of which is clustered and may serve as centromeres. Partial copies of the extrachromosomal ribosomal DNA (rDNA) element are found at the ends of each chromosome, suggesting a novel telomere structure and the use of a common mechanism to maintain both the rDNA and chromosomal termini. A proteome-based phylogeny shows that the amoebozoa diverged from the animal-fungal lineage after the plant-animal split, but Dictyostelium seems to have retained more of the diversity of the ancestral genome than have plants, animals or fungi.

Clostridium botulinum Type F: Isolation from Salmon from the Columbia River.

Clostridium botulinum type F has been isolated from a salmon (Oncorhynchus nerka) taken from the Columbia River. Cultures of this type have been reported only twice before-once the bacterium was isolated.

Biochemical and genetic analysis of the mitochondrial response of yeast to BAX and BCL-X(L).

The BCL-2 family includes both proapoptotic (e.g., BAX and BAK) and antiapoptotic (e.g., BCL-2 and BCL-X(L)) molecules. The cell death-regulating activity of BCL-2 members appears to depend on their ability to modulate mitochondrial function, which may include regulation of the mitochondrial permeability transition pore (PTP). We examined the function of BAX and BCL-X(L) using genetic and biochemical approaches in budding yeast because studies with yeast suggest that BCL-2 family members act upon highly conserved mitochondrial components. In this study we found that in wild-type yeast, BAX induced hyperpolarization of mitochondria, production of reactive oxygen species, growth arrest, and cell death; however, cytochrome c was not released detectably despite the induction of mitochondrial dysfunction. Coexpression of BCL-X(L) prevented all BAX-mediated responses. We also assessed the function of BCL-X(L) and BAX in the same strain of Saccharomyces cerevisiae with deletions of selected mitochondrial proteins that have been implicated in the function of BCL-2 family members. BAX-induced growth arrest was independent of the tested mitochondrial components, including voltage-dependent anion channel (VDAC), the catalytic beta subunit or the delta subunit of the F(0)F(1)-ATP synthase, mitochondrial cyclophilin, cytochrome c, and proteins encoded by the mitochondrial genome as revealed by [rho(0)] cells. In contrast, actual cell killing was dependent upon select mitochondrial components including the beta subunit of ATP synthase and mitochondrial genome-encoded proteins but not VDAC. The BCL-X(L) protection from either BAX-induced growth arrest or cell killing proved to be independent of mitochondrial components. Thus, BAX induces two cellular processes in yeast which can each be abrogated by BCL-X(L): cell arrest, which does not require aspects of mitochondrial biochemistry, and cell killing, which does.

Polymerase chain reaction detection of type D simian retrovirus proviral DNA from infected macaques.

A simple polymerase chain reaction (PCR) approach was developed for detection of Type D simian retrovirus (SRV) serogroup 2 proviral DNA using peripheral blood lymphocytes (PBLs) obtained from infected macaques. PCR primer pairs were developed against serogroup 2 envelope (env) gene sequence, and fidelity of PCR fragment amplification was determined using molecularly cloned SRV serogroup 2 (D2/RHE/OR) DNA, and genomic DNA from Raji cells independently infected with different SRV serogroups. One primer pair exhibiting high fidelity was then utilized for PCR detection of serogroup 2 proviral DNA from PBLs, and from cells sorted into immune cell subpopulations by fluorescent-activated cell sorting (FACS). Env PCR fragments were readily detected from as few as 10(4) PBLs or immune cell subpopulations. In addition, highly specific PCR primers against serogroups 1 and 3 were utilized to detect proviral DNA from Raji cells infected with SRV serogroups. In all cases, primers designed to amplify serogroups 1, 2, and 3 proviral DNA were specific for their intended serogroup. This primer information and development of a PCR approach for detection of specific SRV proviral DNA will be of potential utility as a rapid surveillance tool in monitoring type D simian retrovirus infection within Asian macaque colonies.

The viral diseases of fish: a review through 1978. Part 1: Diseases of proven viral etiology.

In this review, a survey is made of the published literature on the viral diseases of fish available up to and including the year 1978. It is divided into two main sections. Part 1 describes 11 diseases where a virus has been isolated and proven to be the causative agent. Part 2 discusses 16 diseases where there is reason to suspect viral etiology because of evidence deriving from electron microscopy or transmission experiments with bacteria-free filtrates of homogenates of diseased tissue, but where final proof of a causative relationship is lacking. The review attempts to provide the most significant information on the disease process itself, in most cases including external signs, fish species susceptible, pathology, geographic distribution, existence of carriers, methods of transmission, and control. It also gives the most recent and significant data concerning the nature of the causative virus, including its cultural, biological, and physicochemical properties, where such information is available.

Incidence of Clostridium botulinum type E in salmon and other marine fish in the Pacific Northwest.

Salmon, sole, cod, oysters, clams, and crabs from ocean waters along the coast of Oregon and Washington were examined for the presence of Clostridium botulinum type E. The organism was detected by identification of the type E toxin in enrichment cultures of the viscera of individual fish. Of 369 salmon specimens, 48 yielded cultures containing toxin lethal to mice, and almost half of the toxic cultures were shown to contain botulinal toxin, chiefly type E. Eighteen of 113 sole and cod specimens, 4 of 22 Dungeness crab specimens, 5 of 16 oyster specimens, and 27 of 115 clam specimens gave rise to cultures containing botulinal toxin which was usually type E, although types A and B were occasionally encountered.

Promoter-activated expression of nerve growth factor for treatment of neurodegenerative diseases.

Genetic transfer approaches have received recent consideration as potential treatment modalities for human central and peripheral nervous system (CNS and PNS, respectively) neurodegenerative disorders, including Parkinson's disease, Alzheimer's disease, and amyotrophic lateral sclerosis. Transplantation of genetically modified cells into the brain represents a promising strategy for the delivery and expression of specific neurotrophic factors, neurotransmitter-synthesizing enzymes, and cellular regulatory proteins for intervention in neurodegenerative diseases. The use of specific regulatable promoters may also provide potential control of gene expression required for dose-specific or time-specific therapeutic strategies. In this article, we review the potential use of activated promoters in ex vivo systems for the potential genetic therapy of neurodegenerative disorders, and then describe our own studies using the zinc-inducible metallothionein promoter for the regulated expression of nerve growth factor (NGF) in rodent brain transplants.

Molecular cloning and cell-specific growth characterization of polymorphic variants of type D serogroup 2 simian retroviruses.

Simian retroviruses (SRVs), the etiological agent of a spontaneous Simian acquired immunodeficiency syndrome, endemically infects large percentages of Asian macaques housed in biomedical research colonies and severely compromises the effective use of these species as a viable research animal. We recently described the molecular cloning of a serogroup 2 SRV, D2/RHE/OR, which causes mild immunosuppression in rhesus macaques. A restriction site variant, D2/RHE/OR/V1, has also been recovered from severely ill animals endemically infected with D2/RHE/OR. We now report the complete nucleotide sequences of D2/RHE/OR and D2/RHE/OR/V1. Both infectious molecular clones retain the genetic structure typical of type D SRVs (5' LTR-gag-prt-pol-env-3'LTR) and encode identically sized 8105-bp proviruses. D2/RHE/OR and D2/RHE/OR/V1 are 99.3% similar at the amino acid level, exhibiting only 17 residue differences, of which 10 are located in the envelope glycoproteins. The molecular clones and reciprocal chimeric viruses were used to assess the contribution of different genetic domains to virus infectivity in a T cell infection assay. These experiments indicate that D2/RHE/OR has a reduced ability to infect specific T cell lines, especially Hut-78 and MT-4 cells, and that the envelope gene is not the sole determinant of in vitro tropism.

Simian AIDS type D serogroup 2 retrovirus: isolation of an infectious molecular clone and sequence analyses of its envelope glycoprotein gene and 3' long terminal repeat.

We describe the molecular cloning of a serogroup 2 simian retrovirus (SRV; D2/RHE/OR) and present the sequence of its envelope (env) glycoprotein gene and 3' long terminal repeat region. This report documents the first infectious molecular clone of a serogroup 2 SRV and provides env sequence verification of genetic diversity among serogroup 2 SRV isolates.