Abstract
During biotrophy, filamentous pathogens such as the rice blast fungus Magnaporthe oryzae deliver effector proteins into live host cells to facilitate colonization. We describe three complementary assays for visualizing M. oryzae effector translocation into the rice cytoplasm and cell-to-cell movement during infection. Our assays make use of live-cell confocal imaging of optically clear rice sheath cells infected with transgenic strains of M. oryzae that express the fluorescent protein-tagged effector known as PWL2. We highlight several important considerations for the analysis of effector translocation and movement dynamics during infection of host plants.
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References
Giraldo MC, Valent B (2013) Filamentous plant pathogen effectors in action. Nat Rev Micro 11:800–814
Khang CH, Berruyer R, Giraldo MC, Kankanala P, Park S-Y, Czymmek K, Kang S, Valent B (2010) Translocation of Magnaporthe oryzae effectors into rice cells and their subsequent cell-to-cell movement. Plant Cell 22:1388–1403
Presti LL, Lanver D, Schweizer G, Tanaka S, Liang L, Tollot M, Zuccaro A, Reissmann S, Kahmann R (2015) Fungal effectors and plant susceptibility. Annu Rev Plant Biol 66:513–545
Giraldo MC, Dagdas YF, Gupta YK, Mentlak TA, Yi M, Martinez-Rocha AL, Saitoh H, Terauchi R, Talbot NJ, Valent B (2013) Two distinct secretion systems facilitate tissue invasion by the rice blast fungus Magnaporthe oryzae. Nat Commun 4:1996. https://doi.org/10.1038/ncomms2996
Mentlak TA, Kombrink A, Shinya T, Ryder LS, Otomo I, Saitoh H, Terauchi R, Nishizawa Y, Shibuya N, Thomma BP, Talbot NJ (2012) Effector-mediated suppression of chitin-triggered immunity by Magnaporthe oryzae is necessary for rice blast disease. Plant Cell 24:322–335
Mosquera G, Giraldo MC, Khang CH, Coughlan S, Valent B (2009) Interaction transcriptome analysis identifies Magnaporthe oryzae BAS1-4 as biotrophy-associated secreted proteins in rice blast disease. Plant Cell 21:1273–1290
Park C-H, Chen S, Shirsekar G, Zhou B, Khang CH, Songkumarn P, Afzal AJ, Ning Y, Wang R, Bellizzi M, Valent B, Wang G-L (2012) The Magnaporthe oryzae effector AvrPiz-t targets the RING E3 ubiquitin ligase APIP6 to suppress pathogen-associated molecular pattern–triggered immunity in rice. Plant Cell 24:4748–4762
Sharpee W, Oh Y, Yi M, Franck W, Eyre A, Okagaki LH, Valent B, Dean RA (2017) Identification and characterization of suppressors of plant cell death (SPD) effectors from Magnaporthe oryzae. Mol Plant Pathol 18:850–863
Kleemann J, Rincon-Rivera LJ, Takahara H, Neumann U, Ver Loren van Themaat E et al (2012) Sequential delivery of host-induced virulence effectors by appressoria and intracellular hyphae of the phytopathogen Colletotrichum higginsianum. PLoS Pathog 8:e1002643. https://doi.org/10.1371/journal.ppat.1002643
Jones K, Zhu J, Jenkinson CB, Kim DW, Khang CH (2017) Disruption of the interfacial membrane leads to Magnaporthe oryzae effector re-location and lifestyle switch during rice blast disease. bioRxiv. https://doi.org/10.1101/177147
Mochizuki S, Minami E, Nishizawa Y (2015) Live-cell imaging of rice cytological changes reveals the importance of host vacuole maintenance for biotrophic invasion by blast fungus, Magnaporthe oryzae. MicrobiologyOpen 4:952–966
Valent B, Farrall L, Chumley FG (1991) Magnaporthe grisea genes for pathogenicity and virulence identified through a series of backcrosses. Genetics 127:87–101
Mullins ED, Chen X, Romaine P, Raina R, Geiser DM, Kang S (2001) Agrobacterium-mediated transformation of Fusarium oxysporum: an efficient tool for insertional mutagenesis and gene transfer. Phytopathology 91:173–180
Khang CH, Park S-Y, Lee Y-H, Kang S (2005) A dual selection based, targeted gene replacement tool for Magnaporthe grisea and Fusarium oxysporum. Fungal Genet Biol 42:483–492
Kankanala P, Czymmek K, Valent B (2007) Roles for rice membrane dynamics and plasmodesmata during biotrophic invasion by the blast fungus. Plant Cell 19:706–724
Jones K, Kim DW, Park JS, Khang CH (2016) Live-cell fluorescence imaging to investigate the dynamics of plant cell death during infection by the rice blast fungus Magnaporthe oryzae. BMC Plant Biol 16:1–8
Shipman EN, Jones K, Jenkinson CB, Kim DW, Zhu J, Khang CH (2017) Nuclear and structural dynamics during the establishment of a specialized effector-secreting cell by Magnaporthe oryzae in living rice cells. BMC Cell Biol 18:11. https://doi.org/10.1186/s12860-017-0126-z
Oparka KJ (1994) Plasmolysis: new insights into an old process. New Phytol 126:571–591
Acknowledgments
We acknowledge the assistance of the Biomedical Microscopy Core at the University of Georgia with imaging using a Zeiss LSM 710 confocal microscope. This work was supported by the Agriculture and Food Research Initiative competitive grants program, Award number 2014-67013-21717 from the USDA National Institute of Food and Agriculture.
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Jones, K., Khang, C.H. (2018). Visualizing the Movement of Magnaporthe oryzae Effector Proteins in Rice Cells During Infection. In: Ma, W., Wolpert, T. (eds) Plant Pathogenic Fungi and Oomycetes. Methods in Molecular Biology, vol 1848. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-8724-5_9
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DOI: https://doi.org/10.1007/978-1-4939-8724-5_9
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