Homologous Expression and Quantitative Analysis of T3SS-Dependent Secretion of TAP-Tagged XoAvrBs2 in Xanthomonas oryzae pv. oryzae Induced by Rice Leaf Extract원문보기
Kim, S.H.
(Department of Advanced Technology Fusion, Konkuk University)
,
Lee, S.E.
(Monsanto Korea, Inc., Vegetable Seeds Division)
,
Hong, M.K.
(Department of Advanced Technology Fusion, Konkuk University)
,
Song, N.H.
(Department of Green Life Science, College of Convergence, Sangmyung University)
,
Yoon, B.
(Department of Life Science, College of Natural Science, Kyonggi University)
,
Viet, P.T.
(Department of Advanced Technology Fusion, Konkuk University)
,
Ahn, Y.J.
(Department of Life Science, College of Natural Science, Kyonggi University)
,
Lee, B.M.
(Genomics Division, National Academy of Agricultural Science (NAAS), Rural Development Administration (RDA))
,
Jung, J.W.
(Department of Molecular Biotechnology, Konkuk University)
,
Kim, K.P.
(Department of Molecular Biotechnology, Konkuk University)
,
Han, Y.S.
(Department of Advanced Technology Fusion, Konkuk University)
,
Kim, J.G.
(Genomics Division, National Academy of Agricultural Science (NAAS), Rural Development Administration (RDA))
,
Kang, L.W.
(Department of Advanced Technology Fusion, Konkuk University)
Xanthomonas oryzae pv. oryzae (Xoo) produces a putative effector, XoAvrBs2. We expressed XoAvrBs2 homologously in Xoo with a TAP-tag at the C-terminus to enable quantitative analysis of protein expression and secretion. Addition of rice leaf extracts from both Xoo-sensitive and Xoo-resistant rice cu...
Xanthomonas oryzae pv. oryzae (Xoo) produces a putative effector, XoAvrBs2. We expressed XoAvrBs2 homologously in Xoo with a TAP-tag at the C-terminus to enable quantitative analysis of protein expression and secretion. Addition of rice leaf extracts from both Xoo-sensitive and Xoo-resistant rice cultivars to the Xoo cells induced expression of the XoAvrBs2 gene at the transcriptional and translational levels, and also stimulated a remarkable amount of XoAvrBs2 secretion into the medium. In a T3SS-defective Xoo mutant strain, secretion of the TAPtagged XoAvrBs2 was blocked. Thus, we elucidated the transcriptional and translational expressions of the XoAvrBs2 gene in Xoo was induced in vitro by the interaction with rice and the induced secretion of XoAvrBs2 was T3SSdependent. It is the first report to measure the homologous expression and secretion of XoAvrBs2 in vitro by rice leaf extract. Our system for the quantitative analysis of effector protein expression and secretion could be generally used for the study of host-pathogen interactions.
Xanthomonas oryzae pv. oryzae (Xoo) produces a putative effector, XoAvrBs2. We expressed XoAvrBs2 homologously in Xoo with a TAP-tag at the C-terminus to enable quantitative analysis of protein expression and secretion. Addition of rice leaf extracts from both Xoo-sensitive and Xoo-resistant rice cultivars to the Xoo cells induced expression of the XoAvrBs2 gene at the transcriptional and translational levels, and also stimulated a remarkable amount of XoAvrBs2 secretion into the medium. In a T3SS-defective Xoo mutant strain, secretion of the TAPtagged XoAvrBs2 was blocked. Thus, we elucidated the transcriptional and translational expressions of the XoAvrBs2 gene in Xoo was induced in vitro by the interaction with rice and the induced secretion of XoAvrBs2 was T3SSdependent. It is the first report to measure the homologous expression and secretion of XoAvrBs2 in vitro by rice leaf extract. Our system for the quantitative analysis of effector protein expression and secretion could be generally used for the study of host-pathogen interactions.
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제안 방법
This is the first report to measure homologous expression and secretion of a putative effector XoAvrBs2 quantitatively in vitro, and the developed system can be used to study the detailed mechanism underlying the Xoo-rice interactions in order to express and secrete XoAvrBs2.
대상 데이터
Xoo strain KACC13129 (ΔhrcC) was provided by KACC (Korean Agricultural Culture Collection).
성능/효과
Our result showed that the treatment of RLX into Xoo cells was enough to induce XoAvrBs2-TAP secretion out of the Xoo cells. Homologously expressed and secreted XoAvrBs2-TAP from the transformed Xoo was measured quantitatively using dot blots, and transcriptional and translational expressions of the XoAvrBs2 gene were increased, and approximately 96% of total expressed XoAvrBs2-TAP was secreted out of Xoo in 20 h. In Xcv, the translocation of Xanthomonas outer protein (Xop) effectors to host pepper cells was confirmed in vivo at 8 h after Xcv’s inoculation to pepper leaves, and the quantity of the translocated Xop effectors was approximately 10- to 1,000-fold higher compared with control [12].
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