Dataset Information

Uncoupling of the functions of the Arabidopsis VIP1 protein in transient and stable plant genetic transformation by Agrobacterium.

ABSTRACT: Agrobacterium-mediated genetic transformation of plants, a unique example of transkingdom DNA transfer, requires the presence of several proteins encoded by the host cell. One such cellular factor is VIP1, an Arabidopsis protein proposed to interact with and facilitate import of the bacterial DNA-protein transport (T) complexes into the plant cell nucleus. Thus, VIP1 is required for transient expression of the bacterial DNA, an early step in the transformation process. However, the role of VIP1 in subsequent transformation events leading to the stable expression of bacterial DNA was unexplored. Here, we used reverse genetics to dissect VIP1 functionally and demonstrate its involvement in the stable genetic transformation of Arabidopsis plants by Agrobacterium. Our data indicate that the ability of VIP1 to interact with the VirE2 protein component of the T-complex and localize to the cell nucleus is sufficient for transient genetic transformation, whereas its ability to form homomultimers and interact with the host cell H2A histone in planta is required for tumorigenesis and, by implication, stable genetic transformation.

SUBMITTER: Li J

PROVIDER: S-EPMC556277 | biostudies-literature | 2005 Apr

REPOSITORIES: biostudies-literature

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Uncoupling of the functions of the Arabidopsis VIP1 protein in transient and stable plant genetic transformation by Agrobacterium.

Li Jianxiong J Krichevsky Alexander A Vaidya Manjusha M Tzfira Tzvi T Citovsky Vitaly V

Proceedings of the National Academy of Sciences of the United States of America 20050411 16

Agrobacterium-mediated genetic transformation of plants, a unique example of transkingdom DNA transfer, requires the presence of several proteins encoded by the host cell. One such cellular factor is VIP1, an Arabidopsis protein proposed to interact with and facilitate import of the bacterial DNA-protein transport (T) complexes into the plant cell nucleus. Thus, VIP1 is required for transient expression of the bacterial DNA, an early step in the transformation process. However, the role of VIP1 ...[more]

PMID: 15824315

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Project description:BACKGROUND:Plant genome sequencing has resulted in the identification of a large number of uncharacterized genes. To investigate these unknown gene functions, several transient transformation systems have been developed as quick and convenient alternatives to the lengthy transgenic assay. These transient assays include biolistic bombardment, protoplast transfection and Agrobacterium-mediated transient transformation, each having advantages and disadvantages depending on the research purposes. RESULTS:We present a novel transient assay based on cocultivation of young Arabidopsis (Arabidopsis thaliana) seedlings with Agrobacterium tumefaciens in the presence of a surfactant which does not require any dedicated equipment and can be carried out within one week from sowing seeds to protein analysis. This Fast Agro-mediated Seedling Transformation (FAST) was used successfully to express a wide variety of constructs driven by different promoters in Arabidopsis seedling cotyledons (but not roots) in diverse genetic backgrounds. Localizations of three previously uncharacterized proteins were identified by cotransformation with fluorescent organelle markers. The FAST procedure requires minimal handling of seedlings and was also adaptable for use in 96-well plates. The high transformation efficiency of the FAST procedure enabled protein detection from eight transformed seedlings by immunoblotting. Protein-protein interaction, in this case HY5 homodimerization, was readily detected in FAST-treated seedlings with Förster resonance energy transfer and bimolecular fluorescence complementation techniques. Initial tests demonstrated that the FAST procedure can also be applied to other dicot and monocot species, including tobacco, tomato, rice and switchgrass. CONCLUSION:The FAST system provides a rapid, efficient and economical assay of gene function in intact plants with minimal manual handling and without dedicated device. This method is potentially ideal for future automated high-throughput analysis.

Dataset Information

Uncoupling of the functions of the Arabidopsis VIP1 protein in transient and stable plant genetic transformation by Agrobacterium.

Publications

Uncoupling of the functions of the Arabidopsis VIP1 protein in transient and stable plant genetic transformation by Agrobacterium.

Similar Datasets

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