Michael J. Campbell

PANTHER: a browsable database of gene products organized by biological function, using curated protein family and subfamily classification

The PANTHER database was designed for high-throughput analysis of protein sequences. One of the key features is a simplified ontology of protein function, which allows browsing of the database by biological functions. Biologist curators have associated the ontology terms with groups of protein sequences rather than individual sequences. Statistical models (Hidden Markov Models, or HMMs) are built from each of these groups. The advantage of this approach is that new sequences can be automatically classified as they become available. To ensure accurate functional classification, HMMs are constructed not only for families, but also for functionally distinct subfamilies. Multiple sequence alignments and phylogenetic trees, including curator-assigned information, are available for each family. The current version of the PANTHER database includes training sequences from all organisms in the GenBank non-redundant protein database, and the HMMs have been used to classify gene products across the entire genomes of human, and Drosophila melanogaster. The ontology terms and protein families and subfamilies, as well as Drosophila gene c;assifications, can be browsed and searched for free. Due to outstanding contractual obligations, access to human gene classifications and to protein family trees and multiple sequence alignments will temporarily require a nominal registration fee. PANTHER is publicly available on the web at http://panther.celera.com.

Transcriptional regulation and function during the human cell cycle

We report here the transcriptional profiling of the cell cycle on a genome-wide scale in human fibroblasts. We identified approximately 700 genes that display transcriptional fluctuation with a periodicity consistent with that of the cell cycle. Systematic analysis of these genes revealed functional organization within groups of coregulated transcripts. A diverse set of cytoskeletal reorganization genes exhibit cell-cycle–dependent regulation, indicating that biological pathways are redirected for the execution of cell division. Many genes involved in cell motility and remodeling of the extracellular matrix are expressed predominantly in M phase, indicating a mechanism for balancing proliferative and invasive cellular behavior. Transcripts upregulated during S phase displayed extensive overlap with genes induced by DNA damage; cell-cycle–regulated transcripts may therefore constitute coherent programs used in response to external stimuli. Our data also provide clues to biological function for hundreds of previously uncharacterized human genes.

Applications for protein sequence–function evolution data: mRNA/protein expression analysis and coding SNP scoring tools

The vast amount of protein sequence data now available, together with accumulating experimental knowledge of protein function, enables modeling of protein sequence and function evolution. The PANTHER database was designed to model evolutionary sequence–function relationships on a large scale. There are a number of applications for these data, and we have implemented web services that address three of them. The first is a protein classification service. Proteins can be classified, using only their amino acid sequences, to evolutionary groups at both the family and subfamily levels. Specific subfamilies, and often families, are further classified when possible according to their functions, including molecular function and the biological processes and pathways they participate in. The second application, then, is an expression data analysis service, where functional classification information can help find biological patterns in the data obtained from genome-wide experiments. The third application is a coding single-nucleotide polymorphism scoring service. In this case, information about evolutionarily related proteins is used to assess the likelihood of a deleterious effect on protein function arising from a single substitution at a specific amino acid position in the protein. All three web services are available at .

ATP-mediated conformational changes in the RecA filament.

The crystal structure of the E. coli RecA protein was solved more than 10 years ago, but it has provided limited insight into the mechanism of homologous genetic recombination. Using electron microscopy, we have reconstructed five different states of RecA-DNA filaments. The C-terminal lobe of the RecA protein is modulated by the state of the distantly bound nucleotide, and this allosteric coupling can explain how mutations and truncations of this C-terminal lobe enhance RecAs activity. A model generated from these reconstructions shows that the nucleotide binding core is substantially rotated from its position in the RecA crystal filament, resulting in ATP binding between subunits. This simple rotation can explain the large cooperativity in ATP hydrolysis observed for RecA-DNA filaments.

Transcription, genomes, function

Large-scale studies of mRNA expression have displayed the unusual ability to both challenge traditional biological paradigms and enjoy rapid adoption among a wide range of researchers. The proliferating applications of this technology are poised to exert heavy influence on the very way biologists conceptualize problems and ask questions in the post-genome era.