Genomic position affects the expression of tobacco mosaic virus movement and coat protein genes.

Abstract

Alterations in the genomic position of the tobacco mosaic virus (TMV) genes encoding the 30-kDa cellto-cell movement protein or the coat protein greatly affected their expression. Higher production of 30-kDa protein was correlated with increased proximity of the gene to the viral 3' terminus. A mutant placing the 30-kDa open reading frame 207 nucleotides nearer the 3' terminus produced at least 4 times the wild-type TMV 30-kDa protein level, while a mutant placing the 30-kDa open reading frame 470 nucleotides doser to the 3' terminus produced at least 8 times the wild-type TMV 30-kDa protein level. Increases in 30-kDa protein production were not correlated with the subgenomic mRNA promoter (SGP) controlling the 30-kDa gene, since mutants with either the native 30-kDa SGP or the coat protein SGP in front of the 30-kDa gene produced similar levels of 30-kDa protein. Lack of coat protein did not affect 30-kDa protein expression, since a mutant with the coat protein start codon removed did not produce increased amounts of 30-kDa protein. Effects of gene positioning on coat protein expression were examined by using a mutant containing two different tandemly positioned tobamovirus (TMV and Odontoglossum ringspot virus) coat protein genes. Only coat protein expressed from the gene positioned nearest the 3' viral terminus was detected. Analysis of 30-kDa and coat protein subgenomic mRNAs revealed no proportional increase in the levels of mRNA relative to the observed levels of 30-kDa and coat proteins. This suggests that a translational mechanism is primarily responsible for the observed effect of genomic position on expression of 30-kDa movement and coat protein genes.Positive-sense RNA viruses have evolved numerous strategies for gene regulation. One common strategy is the use of subgenomic mRNAs, transcribed from genomic RNA, for the expression of internal open reading frames (ORFs). In this manner the expression of multiple genes residing on a common genomic RNA can be independently regulated. One such virus that utilizes this strategy is the type member of the tobamovirus group, tobacco mosaic virus (TMV).The genome of TMV resides on a single strand of positivesense RNA, 6395 nucleotides (nt) in length, and encodes at least four proteins (1). The TMV genome is organized such that two 5'-coterminal ORFs encoding 126-kDa and 18&-kDa replicase proteins are translated from the genomic RNA, while an internal ORF encoding the 30-kDa cell-to-cellmovement protein and a 3'-proximal ORF encoding the coat protein (structural) are translated from respective subgenomic mRNAs (2-5). A third subgenomic mRNA has been detected for an additional internal ORF, within the 183-kDa ORF, encoding a putative 54-kDa protein that has not yet been detected in infected plants (6). The TMV subgenomic mRNAs are transcribed from negative-sense genomic RNA and share a common 3' terminus.Of particular interest is that genes of TMV expressed via subgenomic mRNAs are independently regulated, both quantitatively and temporally. The 30-kDa pro...

Enhancement or Attenuation of Disease by Deletion of Genes from Citrus Tristeza Virus

“…One possibility was that the deletions caused an increase in expression of other CTV genes, which in turn induced the stem pitting. A common rule of expression of genes of viruses that express multiple genes via subgenomic RNAs is that genes positioned nearer the 3= terminus tend to be produced in larger amounts, and movement of the genes closer to the 3= terminus by deletion of intervening genes increases levels of expression (5). CTV follows this pattern.…”

Degradation of Tobacco Mosaic Virus Movement Protein by the 26S Proteasome

“…Protoplasts infected with TMV-⌬C and treated with MG115 showed a pattern similar to that for cells infected with wild-type TMV, although significantly more MP was observed in cells infected with TMV-⌬C than in cells infected with wild-type TMV. It was previously shown that TMV lacking the CP produces more MP than does wild-type virus (8). When cells infected with a virus lacking a functional MP were treated with MG115, no MP was detected.…”

Replication of tobacco mosaic virus RNA

“…Similarly, there was little di¡erence in the amounts of subgenomic mRNAs produced with constructs in which promoters were placed at di¡erent positions in the RNA. The high level of coat protein produced appears to be controlled mainly at the translational level (Culver et al 1993). Whether the same polymerase synthesizes genomic-length and subgenomic RNAs is not known.…”