Premium
Application of phi LOV 2.1 as a fluorescent marker for visualization of Agrobacterium effector protein translocation
Author(s) -
Roushan Mohammad Reza,
Zeeuw Milou A. M.,
Hooykaas Paul J. J.,
Heusden Gerard Paul H.
Publication year - 2018
Publication title -
the plant journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.058
H-Index - 269
eISSN - 1365-313X
pISSN - 0960-7412
DOI - 10.1111/tpj.14060
Subject(s) - effector , green fluorescent protein , agrobacterium tumefaciens , biology , agrobacterium , microbiology and biotechnology , chromosomal translocation , arabidopsis , secretion , arabidopsis thaliana , fusion protein , transgene , plant cell , nicotiana tabacum , genetics , mutant , gene , biochemistry , recombinant dna
Summary Agrobacterium tumefaciens can genetically transform plants by translocating a piece of oncogenic DNA , called T‐ DNA , into host cells. Transfer is mediated by a type IV secretion system (T4 SS ). Besides the T‐ DNA, which is transferred in a single‐stranded form and at its 5′ end covalently bound to VirD2, several other effector proteins (VirE2, VirE3, VirD5, and VirF) are translocated into the host cells. The fate and function of the translocated proteins inside the host cell are only partly known. Therefore, several studies were conducted to visualize the translocation of the VirE2 protein. As GFP ‐tagged effector proteins are unable to pass the T4 SS , other approaches like the split GFP system were used, but these require specific transgenic recipient cells expressing the complementary part of GFP . Here, we investigated whether use can be made of the photostable variant of LOV , phi LOV 2.1, to visualize effector protein translocation from Agrobacterium to non‐transgenic yeast and plant cells. We were able to visualize the translocation of all five effector proteins, both to yeast cells, and to cells in Nicotiana tabacum leaves and Arabidopsis thaliana roots. Clear signals were obtained that are easily distinguishable from the background, even in cases in which by comparison the split GFP system did not generate a signal.