Random-set methods identify distinct aspects of the enrichment signal in gene-set analysis

A prespecified set of genes may be enriched, to varying degrees, for genes that have altered expression levels relative to two or more states of a cell. Knowing the enrichment of gene sets defined by functional categories, such as gene ontology (GO) annotations, is valuable for analyzing the biological signals in microarray expression data. A common approach to measuring enrichment is by cross-classifying genes according to membership in a functional category and membership on a selected list of significantly altered genes. A small Fisher's exact test $p$-value, for example, in this $2\times2$ table is indicative of enrichment. Other category analysis methods retain the quantitative gene-level scores and measure significance by referring a category-level statistic to a permutation distribution associated with the original differential expression problem. We describe a class of random-set scoring methods that measure distinct components of the enrichment signal. The class includes Fisher's test based on selected genes and also tests that average gene-level evidence across the category. Averaging and selection methods are compared empirically using Affymetrix data on expression in nasopharyngeal cancer tissue, and theoretically using a location model of differential expression. We find that each method has a domain of superiority in the state space of enrichment problems, and that both methods have benefits in practice. Our analysis also addresses two problems related to multiple-category inference, namely, that equally enriched categories are not detected with equal probability if they are of different sizes, and also that there is dependence among category statistics owing to shared genes. Random-set enrichment calculations do not require Monte Carlo for implementation. They are made available in the R package allez.Comment: Published at http://dx.doi.org/10.1214/07-AOAS104 in the Annals of Applied Statistics (http://www.imstat.org/aoas/) by the Institute of Mathematical Statistics (http://www.imstat.org

Sindbis virus proteins nsP1 and nsP2 contain homology to nonstructural proteins from several RNA plant viruses

Although the genetic organization of tobacco mosaic virus (TMV) differs considerably from that of the tripartite viruses (alfalfa mosaic virus [AlMV] and brome mosaic virus [BMV]), all of these RNA plant viruses share three domains of homology among their nonstructural proteins. One such domain, common to the AlMV and BMV 2a proteins and the readthrough portion of TMV p183, is also homologous to the readthrough protein nsP4 of Sindbis virus (Haseloff et al., Proc. Natl. Acad. Sci. U.S.A. 81:4358-4362, 1984). Two more domains are conserved among the AlMV and BMV 1a proteins and TMV p126. We show here that these domains have homology with portions of the Sindbis proteins nsP1 and nsP2, respectively. These results strengthen the view that the four viruses share mechanistic similarities in their replication strategies and may be evolutionarily related. These results also suggest that either the AlMV 1a, BMV 1a, and TMV p126 proteins are multifunctional or Sindbis proteins nsP1 and nsP2 function together as subunits in a single complex

United States Patent, Number: 5,633,447: PLANT TISSUE COMPRISING A SUBGENOMIC PROMOTER

A subgenomic promoter of a positive strand RNA virus is disclosed which directs the amplified expression of a structural gene in plant tissue. The core region and an upstream activating domain of the subgenomic promoter are identified. This promoter can be utilized in a modified virus. or in an appropriate engineered recombinant DNA derivative which may be chromosomally integrated or maintained as an episome in transformed cells

United States Patent 5,500,360: RNA TRANSFORMATION VECTOR

A + strand RNA viral transformation of host organisms with foreign RNA, and expression of said foreign RNA. The foreign RNA is inserted into an infective RNA viral segment containing replication elements, and allowed to infect the host organism. The invention is exemplified utilizing brome mosaic RNA modified to contain a gene coding for chloramphenicol acetyl transferase (CAT) in the transformation of barley protoplasts

Intermolecular RNA Recombination Occurs at Different Frequencies in Alternate Forms of Brome Mosaic Virus RNA Replication Compartments

Positive-strand RNA viruses replicate their genomes in membrane-bound replication compartments. Brome mosaic virus (BMV) replicates in vesicular invaginations of the endoplasmic reticulum membrane. BMV has served as a productive model system to study processes like virus-host interactions, RNA replication and recombination. Here we present multiple lines of evidence showing that the structure of the viral RNA replication compartments plays a fundamental role and that recruitment of parental RNAs to a common replication compartment is a limiting step in intermolecular RNA recombination. We show that a previously defined requirement for an RNA recruitment element on both parental RNAs is not to function as a preferred crossover site, but in order for individual RNAs to be recruited into the replication compartments. Moreover, modulating the form of the replication compartments from spherular vesicles (spherules) to more expansive membrane layers increased intermolecular RNA recombination frequency by 200- to 1000-fold. We propose that intermolecular RNA recombination requires parental RNAs to be recruited into replication compartments as monomers, and that recruitment of multiple RNAs into a contiguous space is much more common for layers than for spherules. These results could explain differences in recombination frequencies between viruses that replicate in association with smaller spherules versus larger double-membrane vesicles and convoluted membranes

The Host Cell Sulfonation Pathway Contributes to Retroviral Infection at a Step Coincident with Provirus Establishment

The early steps of retrovirus replication leading up to provirus establishment are highly dependent on cellular processes and represent a time when the virus is particularly vulnerable to antivirals and host defense mechanisms. However, the roles played by cellular factors are only partially understood. To identify cellular processes that participate in these critical steps, we employed a high volume screening of insertionally mutagenized somatic cells using a murine leukemia virus (MLV) vector. This approach identified a role for 3′-phosphoadenosine 5′-phosphosulfate synthase 1 (PAPSS1), one of two enzymes that synthesize PAPS, the high energy sulfate donor used in all sulfonation reactions catalyzed by cellular sulfotransferases. The role of the cellular sulfonation pathway was confirmed using chemical inhibitors of PAPS synthases and cellular sulfotransferases. The requirement for sulfonation was mapped to a stage during or shortly after MLV provirus establishment and influenced subsequent gene expression from the viral long terminal repeat (LTR) promoter. Infection of cells by an HIV vector was also shown to be highly dependent on the cellular sulfonation pathway. These studies have uncovered a heretofore unknown regulatory step of retroviral replication, have defined a new biological function for sulfonation in nuclear gene expression, and provide a potentially valuable new target for HIV/AIDS therapy

Establishment of Human Papillomavirus Infection Requires Cell Cycle Progression

Human papillomaviruses (HPVs) are DNA viruses associated with major human cancers. As such there is a strong interest in developing new means, such as vaccines and microbicides, to prevent HPV infections. Developing the latter requires a better understanding of the infectious life cycle of HPVs. The HPV infectious life cycle is closely linked to the differentiation state of the stratified epithelium it infects, with progeny virus only made in the terminally differentiating suprabasal compartment. It has long been recognized that HPV must first establish its infection within the basal layer of stratified epithelium, but why this is the case has not been understood. In part this restriction might reflect specificity of expression of entry receptors. However, this hypothesis could not fully explain the differentiation restriction of HPV infection, since many cell types can be infected with HPVs in monolayer cell culture. Here, we used chemical biology approaches to reveal that cell cycle progression through mitosis is critical for HPV infection. Using infectious HPV16 particles containing the intact viral genome, G1-synchronized human keratinocytes as hosts, and early viral gene expression as a readout for infection, we learned that the recipient cell must enter M phase (mitosis) for HPV infection to take place. Late M phase inhibitors had no effect on infection, whereas G1, S, G2, and early M phase cell cycle inhibitors efficiently prevented infection. We conclude that host cells need to pass through early prophase for successful onset of transcription of the HPV encapsidated genes. These findings provide one reason why HPVs initially establish infections in the basal compartment of stratified epithelia. Only this compartment of the epithelium contains cells progressing through the cell cycle, and therefore it is only in these cells that HPVs can establish their infection. By defining a major condition for cell susceptibility to HPV infection, these results also have potentially important implications for HPV control

Statistical Use of Argonaute Expression and RISC Assembly in microRNA Target Identification

MicroRNAs (miRNAs) posttranscriptionally regulate targeted messenger RNAs (mRNAs) by inducing cleavage or otherwise repressing their translation. We address the problem of detecting m/miRNA targeting relationships in homo sapiens from microarray data by developing statistical models that are motivated by the biological mechanisms used by miRNAs. The focus of our modeling is the construction, activity, and mediation of RNA-induced silencing complexes (RISCs) competent for targeted mRNA cleavage. We demonstrate that regression models accommodating RISC abundance and controlling for other mediating factors fit the expression profiles of known target pairs substantially better than models based on m/miRNA expressions alone, and lead to verifications of computational target pair predictions that are more sensitive than those based on marginal expression levels. Because our models are fully independent of exogenous results from sequence-based computational methods, they are appropriate for use as either a primary or secondary source of information regarding m/miRNA target pair relationships, especially in conjunction with high-throughput expression studies.</p

We Need To Talk: Digital Practices & Ethics in Our Profession

As Student Affairs educators leverage technology for professional practice, we have failed to discuss how our digital lives intersect with our work lives. This competency-based, case study guide is designed to facilitate conversations about expectations and realities of what it means to be a professional online. To help you discuss ways to support digital-ethical professional practice in higher education, we have identified a few scenarios to discuss and develop a positive culture online. We encourage your to start an open dialogue on these issues and identify potential solutions to address unwanted interactions and inappropriate behaviors in professional online networks. Please feel free to bring these case studies back to your campus and/or graduate programs to continue the conversations. This resource is shared with the following Creative Commons license: CC BY-NC-SA 4.0Graduate Students and New Professionals Community of PracticeEducatio