Reference sequence (RefSeq) database at NCBI: current status, taxonomic expansion, and functional annotation.

The RefSeq project at the National Center for Biotechnology Information (NCBI) maintains and curates a publicly available database of annotated genomic, transcript, and protein sequence records (http://www.ncbi.nlm.nih.gov/refseq/). The RefSeq project leverages the data submitted to the International Nucleotide Sequence Database Collaboration (INSDC) against a combination of computation, manual curation, and collaboration to produce a standard set of stable, non-redundant reference sequences. The RefSeq project augments these reference sequences with current knowledge including publications, functional features and informative nomenclature. The database currently represents sequences from more than 55,000 organisms (>4800 viruses, >40,000 prokaryotes and >10,000 eukaryotes; RefSeq release 71), ranging from a single record to complete genomes. This paper summarizes the current status of the viral, prokaryotic, and eukaryotic branches of the RefSeq project, reports on improvements to data access and details efforts to further expand the taxonomic representation of the collection. We also highlight diverse functional curation initiatives that support multiple uses of RefSeq data including taxonomic validation, genome annotation, comparative genomics, and clinical testing. We summarize our approach to utilizing available RNA-Seq and other data types in our manual curation process for vertebrate, plant, and other species, and describe a new direction for prokaryotic genomes and protein name management.

Assembly: a resource for assembled genomes at NCBI.

The NCBI Assembly database (www.ncbi.nlm.nih.gov/assembly/) provides stable accessioning and data tracking for genome assembly data. The model underlying the database can accommodate a range of assembly structures, including sets of unordered contig or scaffold sequences, bacterial genomes consisting of a single complete chromosome, or complex structures such as a human genome with modeled allelic variation. The database provides an assembly accession and version to unambiguously identify the set of sequences that make up a particular version of an assembly, and tracks changes to updated genome assemblies. The Assembly database reports metadata such as assembly names, simple statistical reports of the assembly (number of contigs and scaffolds, contiguity metrics such as contig N50, total sequence length and total gap length) as well as the assembly update history. The Assembly database also tracks the relationship between an assembly submitted to the International Nucleotide Sequence Database Consortium (INSDC) and the assembly represented in the NCBI RefSeq project. Users can find assemblies of interest by querying the Assembly Resource directly or by browsing available assemblies for a particular organism. Links in the Assembly Resource allow users to easily download sequence and annotations for current versions of genome assemblies from the NCBI genomes FTP site.

Distribution of virulence genes in clinical and environmental Vibrio cholerae strains in Bangladesh.

Vibrio cholerae, an environmental organism, is a facultative human pathogen. Here, we report the virulence profiles, comprising 18 genetic markers, of 102 clinical and 692 environmental V. cholerae strains isolated in Bangladesh between March 2004 and January 2006, showing the variability of virulence determinants within the context of public health.

Discovery of novel Vibrio cholerae VSP-II genomic islands using comparative genomic analysis.

This report describes Vibrio seventh pandemic island II (VSP-II) and three novel variants revealed by comparative genomics of 23 Vibrio cholerae strains and their presence among a large and diverse collection of V. cholerae isolates. Three VSP-II variants were reported previously and our results demonstrate the presence of three novel VSP-II in clinical and environmental V. cholerae marked by major deletions and genetic rearrangements. A new VSP-II cluster was found in the seventh pandemic V. cholerae O1 El Tor strain CIRS101, which is dominant (95%) among the recent (2004-2007) seven pandemic V. cholerae O1 El Tor isolates from two endemic sites, but was not found in older strains from the same region. Two other variants were found in V. cholerae TMA21 and RC385, two environmental strains from coastal Brazil and the Chesapeake Bay, respectively, the latter being prevalent among environmental V. cholerae non-O1/non-O139 and Vibrio mimicus. The results of this study indicate that the VSP-II island has undergone significant rearrangement through a complex evolutionary pathway in V. cholerae. Interestingly, one of the new VSP-II revealed the presence of 'old' and 'new' V. cholerae O1 El Tor pandemic clones circulating in some of the areas where cholera is endemic.

Occurrence of the Vibrio cholerae seventh pandemic VSP-I island and a new variant.

Using comparative genomics, we identified a new variant of the Vibrio Seventh Pandemic Island-I (VSP-I). Results of polymerase chain reaction (PCR) screening for both known VSP-I variants indicate that the novel variant is present only in non-O1/non-O139 strains of V. cholerae and Vibrio mimicus. Comparative genomics revealed little sequence divergence in the seventh pandemic VSP-I; however, a second insertion site located on the smaller chromosome was identified. Although the seventh pandemic VSP-I genomic island was detected in all seventh pandemic V. cholerae serogroup O1 and O139 isolates examined in this study, unique genes of the island cannot be used alone as an identifying target, because the seventh pandemic VSP-I was also present in three non-seventh pandemic strains of V. cholerae isolated from Chesapeake Bay. As an alternative, a PCR assay targeting the VC2346 gene was found to be confirmatory for seventh pandemic isolates of V. cholerae.

Anti-apoptotic Bcl-2 Family Proteins Disassemble Ceramide Channels.

Early in mitochondria-mediated apoptosis, the mitochondrial outer membrane becomes permeable to proteins that, when released into the cytosol, initiate the execution phase of apoptosis. Proteins in the Bcl-2 family regulate this permeabilization, but the molecular composition of the mitochondrial outer membrane pore is under debate. We reported previously that at physiologically relevant levels, ceramides form stable channels in mitochondrial outer membranes capable of passing the largest proteins known to exit mitochondria during apoptosis (Siskind, L. J., Kolesnick, R. N., and Colombini, M. (2006) Mitochondrion 6, 118-125). Here we show that Bcl-2 proteins are not required for ceramide to form protein-permeable channels in mitochondrial outer membranes. However, both recombinant human Bcl-x(L) and CED-9, the Caenorhabditis elegans Bcl-2 homologue, disassemble ceramide channels in the mitochondrial outer membranes of isolated mitochondria from rat liver and yeast. Importantly, Bcl-x L and CED-9 disassemble ceramide channels in the defined system of solvent-free planar phospholipid membranes. Thus, ceramide channel disassembly likely results from direct interaction with these anti-apoptotic proteins. Mutants of Bcl-x L act on ceramide channels as expected from their ability to be anti-apoptotic. Thus, ceramide channels may be one mechanism for releasing pro-apoptotic proteins from mitochondria during the induction phase of apoptosis.