The nucleotide sequence of the RNA of tobacco vein mottling virus, a member of the potyvirus group, was determined. The RNA was found to be 9471 residues in length, excluding a 3'-terminal poly(A) tail. The first three AUG codons from the 5'-terminus were followed by in-frame termination codons. The fourth, at position 206, was the beginning of an open reading frame of 9015 residues which could encode a polyprotein of 340 kDa. No other long open reading frames were present in the sequence or its complement. This AUG was present in the sequence AGGCCAUG, which is similar to the consensus initiation sequence shared by most eukaryotic mRNAs. The chemically-determined amino acid compositions of the helper component and coat proteins were similar to those predicted from the nucleotide sequence. Amino acid sequencing of coat protein from which an amino-terminal peptide had been removed allowed exact location of the coat protein cistron. A consensus sequence of V-(R or K)-F-Q was found on the N-terminal sides of proposed cleavage sites for proteolytic processing of the polyprotein.

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Cigarette smoking has been associated with an increase in the severity and prevalence of atherosclerosis in the abdominal aorta. To begin our investigation of this finding, we used an integrated approach combining gene expression profiling, protein analysis, cytokine measurements, and cytotoxicity determinations to examine molecular responses of cultured human aortic and coronary endothelial cells exposed to cigarette smoke condensate (CSC) and nicotine. Exposure of endothelial cells to CSC (30 and 60 microg/mL TPM) for 24 h resulted in minimal cytotoxicity, and the upregulation of genes involved in matrix degradation (MMP-1, MMP-8, and MMP-9), xenobiotic metabolism (HO-1 and CYP1A2), and downregulation of genes involved in cell cycle regulation (including TOP2A, CCNB1, CCNA, CDKN3). Exposure of cells to a high physiological concentration of nicotine resulted in few differentially expressed genes. Immunoblot analysis of proteins selected from genes shown to be differentially regulated by microarray analysis revealed similar responses. Finally, a number of inflammatory cytokines measured in culture media were elevated in response to CSC. Together, these results describe a complex proinflammatory response, possibly mediating the recruitment of leukocytes through cytokine signaling. Additionally, fibrous cap destabilization may be facilitated by matrix metalloproteinase upregulation.

The va gene is used in commercial Burley tobacco cultivars including cv TN 86 to confer resistance to tobacco vein mottling virus (TVMV) and, to some extent, other potyviruses. A naturally occurring strain of TVMV (TVMV-S), which overcomes this resistance, was isolated from TN 86 plants. To investigate the mechanism by which TVMV-S overcomes va gene resistance, a cDNA clone encompassing the complete genome of TVMV-S was produced. Using chimeric transcripts combining regions of TVMV-S and regions of the wild-type strain (TVMV-WT) to which TN 86 is resistant, it was demonstrated that a domain within the VPg protein is responsible for overcoming va resistance in TN 86. DNA sequence analysis revealed six amino acid differences between the two strains of TVMV within this domain. Inclusion of sequences for four of the TVMV-S VPg amino acids was sufficient to confer the resistance-overcoming phenotype to all corresponding transcripts. Coinoculation experiments suggested that the resistance of TN 86 to TVMV-WT was not due to the induction of a general host defense response. The results are compatible with the hypothesis that VPg must assume an appropriate configuration in order to interact with appropriate host components and facilitate systemic virus movement.