DNA microarray analysis of chimpanzee liver during acute resolving hepatitis C virus infection.

Hepatitis C virus (HCV) poses a worldwide health problem in that the majority of individuals exposed to HCV become chronically infected and are predisposed for developing significant liver disease. DNA microarray technology provides an opportunity to survey transcription modulation in the context of an infectious disease and is a particularly attractive approach in characterizing HCV-host interactions, since the mechanisms underlying viral persistence and disease progression are not understood and are difficult to study. Here, we describe the changes in liver gene expression during the course of an acute-resolving HCV infection in a chimpanzee. Clearance of viremia in this animal occurred between weeks 6 and 8, while clearance of residual infected hepatocytes did not occur until 14 weeks postinfection. The most notable changes in gene expression occurred in numerous interferon response genes (including all three classical interferon antiviral pathways) that increased dramatically, some as early as day 2 postinfection. The data suggest a biphasic mechanism of viral clearance dependent on both the innate and adaptive immune responses and provide insight into the response of the liver to a hepatotropic viral infection.

Sp1 and Sp3 regulate expression of the neuronal nicotinic acetylcholine receptor beta4 subunit gene.

Neuronal nicotinic acetylcholine receptors play important roles in signal transduction within the nervous system. The receptors exist in a variety of functionally distinct subtypes that are determined by their subunit structures. The subunits are encoded by 11 genes, alpha2-alpha9 and beta2-beta4. Three of the genes, alpha3, alpha5, and beta4, are tightly clustered, and their encoded proteins make up the predominant receptor subtype in the peripheral nervous system. The tight linkage of the genes suggests there may be a common regulatory mechanism underlying their expression. However, although their expression patterns significantly overlap, they are not identical, indicating that independent regulatory mechanisms must also exist. Our studies have focused upon the gene encoding the beta4 subunit for which we have identified several transcriptional regulatory elements. One of these elements, E2, specifically interacts with the general transcription factor Sp1. Here we show that another member of the Sp family of factors, Sp3, can specifically interact with E2 whereas two other members, Sp2 and Sp4, cannot. Co-transfection experiments indicate that Sp3 can transactivate a beta4 promoter/reporter gene construct and, furthermore, that Sp1 and Sp3 can transactivate the beta4 reporter construct synergistically. The transactivation is dependent upon an intact E2 and may involve direct interactions between Sp1 and Sp3.

Transcriptional regulation of neuronal nicotinic acetylcholine receptor genes. Functional interactions between Sp1 and the rat beta4 subunit gene promoter.

To date, 11 members (alpha2-alpha9 and beta2-beta4) of the neuronal nicotinic acetylcholine receptor gene family have been identified. These genes encode subunits that form distinct receptors with different pharmacological and physiological profiles in temporally and spatially restricted patterns within the nervous system. Distinct molecular mechanisms probably orchestrate the expression of various receptor subtypes, yet little is known of specific transcriptional regulatory elements and their associated factors that are responsible for this segregated pattern of expression. Here we report the identification of an element, in the 5'-flanking region of the rat beta4 subunit gene, containing a CA box that is necessary for beta4 promoter activity in a transiently transfected cholinergic cell line, SN17. This element was shown to interact with a protein(s) in SN17 nuclear extracts that is antigenically related to the transcriptional activator Sp1. Furthermore, co-transfection experiments confirmed that Sp1 can transactivate a beta4 promoter-reporter gene construct, indicating that Sp1 is necessary, at least in part, for transcriptional activation of the beta4 subunit gene.

A novel regulatory element of a nicotinic acetylcholine receptor gene interacts with a DNA binding activity enriched in rat brain.

Nicotinic acetylcholine receptors are ligand-gated ion channels that play a critical role in signal transmission in the nervous system. The genes encoding the various subunits that comprise functional acetylcholine receptors are expressed in distinct temporal and spatial patterns. Studies to understand the molecular mechanisms underlying the differential expression of the receptor subunit genes have led to the identification, in this report, of a 19-base pair cis-acting element that is required for transcriptional activation of the rat beta 4 subunit gene. Screening of computer data bases with the 19-base pair element revealed the sequence to be unique among known transcriptional regulatory elements. Loss of this element resulted in drastically reduced beta 4 promoter activity in transfected cholinergic SN17 cells. Furthermore, this element specifically interacts with nuclear proteins prepared from both SN17 cells and adult rat brain. UV cross-linking experiments indicated the presence, in SN17 nuclear extracts, of a prominent protein species (approximately 50 kDa) that interacts specifically with the 19-base pair element. These results lead us to hypothesize that interactions between the 50-kDa protein and the novel 19-base pair element are necessary for transcriptional activation of the beta 4 subunit gene.