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The TMV movement protein: role of the C-terminal 73 amino acids in subcellular localization and function
Abstract:
The role of the C-terminal one-third of the tobacco mosaic virus (TMV) 30-kDa movement protein (MP) on its subcellular localization and on virus spread was investigated. We have constructed eight cDNAs encoding MPs with variable size deletions from the C-terminal end. Expression of the truncated proteins was verified in recombinant yeast using an antiserum directed to a synthetic peptide corresponding to 21 amino acids near the N-terminal end of the MP. In transgenic tobacco plants, MP from which more than 55 amino acids were deleted no longer accumulated in the cell wall fraction of a cellular extract, where the complete MP accumulates. Dye diffusion studies showed that both unmodified and modified MPs that accumulate in the cell wall fraction are able to alter plasmodesmatal size exclusion limits. Biological function of the modified MPs was tested in the transgenic plants with the TMV thermosensitive mutant Ls1 and a TMV genomic RNA transcript lacking a functional MP. There was a correlation between the cell wall localization of the modified MPs and its ability to potentiate virus spread. The results presented here demonstrate the dispensability of the C-terminal 55 amino acids of the MP in its subcellular localization in tobacco plants and its role in virus movement. Moreover, our results show that a stretch of 19 amino acids (195 to 213) is essential for localization of the MP to the cell wall fraction of plant cells.
Insights
The C-terminal region of the tobacco mosaic virus (TMV) movement protein (MP) is not essential for its function in virus spread. Deleting the final 55 amino acids of the MP affects its cell wall localization and ability to spread the virus.
Area of Science:
- Plant Virology
- Molecular Biology
- Cell Biology
Background:
- The tobacco mosaic virus (TMV) movement protein (MP) facilitates virus spread between plant cells.
- The C-terminal region of TMV MP is implicated in its function, but its specific role in subcellular localization and virus movement requires clarification.
Purpose of the Study:
- To investigate the role of the C-terminal one-third of TMV MP in subcellular localization and virus spread.
- To identify specific amino acid sequences within the C-terminus crucial for MP function.
Main Methods:
- Construction and expression of TMV MP variants with C-terminal deletions in yeast and transgenic tobacco plants.
- Verification of truncated protein expression using specific antisera.
- Analysis of MP subcellular localization in cell wall fractions.
- Dye diffusion assays to assess plasmodesmatal size exclusion limits.
- Evaluation of biological function using TMV mutants in transgenic plants.
Main Results:
- TMV MPs lacking more than 55 C-terminal amino acids did not accumulate in the cell wall fraction.
- MPs localized to the cell wall fraction, whether modified or unmodified, altered plasmodesmatal size exclusion limits.
- A strong correlation was observed between cell wall localization of modified MPs and their ability to potentiate virus spread.
- A 19-amino acid stretch (residues 195-213) was identified as essential for MP localization to the cell wall.
Conclusions:
- The C-terminal 55 amino acids of TMV MP are dispensable for its subcellular localization and role in virus movement in tobacco plants.
- Specific amino acid sequences within the C-terminus, particularly residues 195-213, are critical for targeting the MP to the cell wall, which is linked to its function in potentiating virus spread.