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Central to immune and inflammatory responses are the integrin-mediated adhesive interactions of cells with their extracellular matrix (ECM)-rich environment. Using a comprehensive and quantitative mRNA profiling technique, we analyzed the effect of ECM-induced attachment on monocyte gene expression, its regulation by growth factors, and the integrin specificity of this event. Adhesion of cells to fibronectin resulted in increased expression of a large number of genes, which was strongly potentiated by the presence of growth factors. Adhesion activated both the NF-κB and Jak/STAT pathways of gene transcription and increased expression of genes involved in inflammatory and immune responses, revealing the importance of ECM–integrin interactions in these processes.

Since their development in the early 1990s, differential gene expression (DGE) technologies have been applied to a multitude of biological challenges, both for the purpose of basic biological research and as a valuable tool for the discovery and development of pharmaceuticals. In this review we survey a class of DGE technologies collectively referred to as ‘open’ architecture systems. These technologies are distinct from the ‘closed’ DGE technologies (quantitative PCR, chip technologies), in that no pre-existing biological or sequence information is necessary and they are applicable to any species. Examples of open systems include GeneCalling®, SAGE, TOGA, READS™, and their progenitor DGE technologies, differential display and cDNA representational difference analysis. We review these technologies and summarize a specific application using GeneCalling for novel gene discovery. Additionally, the significance of data management and experimental design in this new age of expression analysis is discussed.

The 60-kDa Ro autoantigen is normally complexed with small cytoplasmic RNAs known as Y RNAs. In Xenopus oocytes, the Ro protein is also complexed with a large class of variant 5S rRNA precursors that are folded incorrectly. Using purified baculovirus-expressed protein, we show that the 60-kDa Ro protein binds directly to both Y RNAs and misfolded 5S rRNA precursors. To understand how the protein recognizes these two distinct classes of RNAs, we investigated the features of Y RNA sequence and structure that are necessary for protein recognition. We identified a truncated Y RNA that is stably bound by the 60-kDa Ro protein. Within this 39-nt RNA is a conserved helix that is proposed to be the binding site for the Ro protein. Mutagenesis of this minimal Y RNA revealed that binding by the 60-kDa Ro protein requires specific base pairs within the conserved helix, a singly bulged nucleotide that disrupts the helix, and a three-nucleotide bulge on the opposing strand. Chemical probing experiments using diethyl pyrocarbonate demonstrated that, in the presence of the two bulges, the major groove of the conserved helix is accessible to protein side chains. These data are consistent with a model in which the Ro protein recognizes specific base pairs in the conserved helix by binding in the major groove of the RNA. Furthermore, experiments in which dimethyl sulfate was used to probe a naked and protein-bound Y RNA revealed that a structural alteration occurs in the RNA upon Ro protein binding.

Plastids present in different tissues may vary morphologically and functionally, despite the fact that all plastids within the same plant contain identical genomes. This is achieved by regulation of expression of the plastid genome by tissue-specific factors, the mechanisms of which are not fully understood. The proton translocating ATP synthase/ATPase is a multisubunit complex composed of nine subunits, six encoded in the plastid and three in the nucleus. We have investigated the tissue-specific expression of the large ATP synthase gene cluster in spinach (Spinacia oleracea). This gene cluster encodes four of the six plastid-encoded ATP synthase genes. Transcript abundance, transcriptional activity, and transcript stability were investigated relative to gene dosage in root plastids and in stem, leaf, and flower chloroplasts. All three of these factors display significant tissue-specific variation. It was intriguing to discover that, although transcript abundance normalized to gene dosage varies in each tissue, transcript abundance as a proportion of the entire plastid RNA population in each tissue is not significantly different. Thus it appears that in these tissues the variation in transcription and stability of transcripts derived from the large ATP synthase gene cluster balances to yield an equivalent proportion of these transcripts in the plastid RNA population. Expression of this gene cluster in photosynthetic as well as non-photosynthetic tissues may facilitate the plasticity of structure and function which is characteristic of plastids.