Agrobacterium VirE2 Protein Modulates Plant Gene Expression and Mediates Transformation From Its Location Outside the Nucleus

Abstract

Agrobacterium effector protein VirE2 is important for plant transformation. VirE2 likely coats transferred DNA (T-DNA) in the plant cell and protects it from degradation. VirE2 localizes to the plant cytoplasm and interacts with several host proteins. Plant-expressed VirE2 can complement a virE2 mutant Agrobacterium strain to support transformation. We investigated whether VirE2 could facilitate transformation from a nuclear location by affixing to it a strong nuclear localization signal (NLS) sequence. Only cytoplasmic-, but not nuclear-localized, VirE2 could stimulate transformation. To investigate the ways VirE2 supports transformation, we generated transgenic Arabidopsis plants containing a virE2 gene under the control of an inducible promoter and performed RNA-seq and proteomic analyses before and after induction. Some differentially expressed plant genes were previously known to facilitate transformation. Knockout mutant lines of some other VirE2 differentially expressed genes showed altered transformation phenotypes. Levels of some proteins known to be important for transformation increased in response to VirE2 induction, but prior to or without induction of their corresponding mRNAs. Overexpression of some other genes whose proteins increased after VirE2 induction resulted in increased transformation susceptibility. We conclude that cytoplasmically localized VirE2 modulates both plant RNA and protein levels to facilitate transformation.

Myosin VIII and XI isoforms interact withAgrobacteriumVirE2 protein and help direct transport from the plasma membrane to the perinuclear region during plant transformation

“…Six days later we assessed transient transformation efficiency by conducting β-glucuronidase (GUS) activity assays using the chromogenic substrate 5-bromo-4-chloro-3-indolyl-β-D-glucuronide (X-gluc). As previously reported (Lapham et al, 2021), induction of the VirE2-Venus transgene in wild-type Col-0 plants complemented the loss of VirE2 function when a virE2 Agrobacterium mutant was used. However, induction of the VirE2-Venus transgene in myosin VIII quadruple mutant plants could not compensate the loss of VirE2 function when a virE2 Agrobacterium mutant was used (Figures 4A and 4B).…”

“…Fewer T-strands can be isolated from plant cells infected by an Agrobacterium virE2 mutant than from cells infected by a wild-type Agrobacterium strain (Yusibov et al, 1994), and the few transformants produced by a virE2 mutant Agrobacterium strain generally harbor large T-DNA deletions, suggesting a protective function of VirE2 on T-strands (Rossi et al, 1996). However, a virE2 mutant Agrobacterium strain can efficiently transform plants expressing VirE2 ( in planta complementation), indicating both the importance of VirE2 in transformation and that the function of VirE2 in transformation occurs within the plant (Citovsky et al, 1992; Lapham et al, 2021). The efficient rescue of an Agrobacterium virE2 mutant by in planta complementation indicates that VirE2 trafficking within a plant cell can be studied using plant lines that produce VirE2, rather than studying VirE2 trafficking after delivery from Agrobacterium .…”

“…(in planta complementation), indicating both the importance of VirE2 in transformation and that the function of VirE2 in transformation occurs within the plant (Citovsky et al, 1992;Lapham et al, 2021). The efficient rescue of an Agrobacterium virE2 mutant by in planta complementation indicates that VirE2 trafficking within a plant cell can be studied using plant lines that produce VirE2, rather than studying VirE2 trafficking after delivery from Agrobacterium.…”

Genotype-Dependent Effect of Silencing of TaCKX1 and TaCKX2 on Phytohormone Crosstalk and Yield-Related Traits in Wheat

“…Regulation of T-DNA expression by DNA methylation induces the host-plant defense response by global changes in plant growth regulation. As recently reported, Agrobacterium VirE2, which coats single-stranded T-DNA molecules, might alter the transcriptome and proteome of the Arabidopsis root system to facilitate transformation [ 42 ]. Moreover, such modifications of DNA methylation and other epigenetic modifications occur during in vitro culture as well [ 43 ].…”

Agrobacterium-mediated gene transfer: recent advancements and layered immunity in plants

“…(Shi et al 2014 ). According to Lapham et al, VirE2 localised in the cytoplasm modulates plant RNAs and genes to promote transformation (Lapham et al 2021 ).…”