Host-induced gene silencing of wheat leaf rust fungus Puccinia triticina pathogenicity genes mediated by the Barley stripe mosaic virus

Host-induced gene silencing of wheat leaf rust fungus Puccinia triticina pathogenicity genes... Rust fungi are devastating plant pathogens and several Puccinia species have a large economic impact on wheat production worldwide. Disease protection, mostly offered by introgressed host-resistance genes, is often race-specific and rapidly overcome by newly-emerging virulent strains. Extensive new genomic resources have identified vital pathogenicity genes but their study is hampered because of the biotrophic life styles of rust fungi. In cereals, Barley stripe mosaic virus (BSMV)-induced RNAi has emerged as a useful tool to study loss-of-function phenotypes of candidate genes. Expression of pathogen-derived gene fragments in this system can be used to obtain in planta-generated silencing of corresponding genes inside biotrophic pathogens, a technique termed host-induced gene silencing (HIGS). Here we test the effectiveness of BSMV-mediated HIGS in the wheat leaf rust fungus Puccinia triticina (Pt) by targeting three predicted pathogenicity genes, a MAPK, a cyclophilin, and a calcineurin regulatory subunit. Inoculation of BSMV RNAi constructs generated fungal gene-specific siRNA molecules in systemic leaves of wheat plant. Subsequent Pt inoculation resulted in a suppressed disease phenotype and a reduction in endogenous transcript levels of the targeted fungal genes indicating translocation of siRNA molecules from host to fungal cells. Efficiency of this host-generated trans-specific RNAi was enhanced by using BSMV silencing vectors defective in coat protein coupled with introducing fungal gene sequences simultaneously in sense and antisense orientation. The disease suppression indicated the likely involvement of these fungal genes in pathogenicity. This study demonstrates that BSMV-mediated in planta-generated RNAi is an effective strategy for functional genomics in rust fungi. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Plant Molecular Biology Springer Journals

Host-induced gene silencing of wheat leaf rust fungus Puccinia triticina pathogenicity genes mediated by the Barley stripe mosaic virus

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Publisher
Springer Netherlands
Copyright
Copyright © 2013 by Her Majesty the Queen in Right of Canada
Subject
Life Sciences; Plant Sciences; Biochemistry, general; Plant Pathology
ISSN
0167-4412
eISSN
1573-5028
D.O.I.
10.1007/s11103-013-0022-7
Publisher site
See Article on Publisher Site

Abstract

Rust fungi are devastating plant pathogens and several Puccinia species have a large economic impact on wheat production worldwide. Disease protection, mostly offered by introgressed host-resistance genes, is often race-specific and rapidly overcome by newly-emerging virulent strains. Extensive new genomic resources have identified vital pathogenicity genes but their study is hampered because of the biotrophic life styles of rust fungi. In cereals, Barley stripe mosaic virus (BSMV)-induced RNAi has emerged as a useful tool to study loss-of-function phenotypes of candidate genes. Expression of pathogen-derived gene fragments in this system can be used to obtain in planta-generated silencing of corresponding genes inside biotrophic pathogens, a technique termed host-induced gene silencing (HIGS). Here we test the effectiveness of BSMV-mediated HIGS in the wheat leaf rust fungus Puccinia triticina (Pt) by targeting three predicted pathogenicity genes, a MAPK, a cyclophilin, and a calcineurin regulatory subunit. Inoculation of BSMV RNAi constructs generated fungal gene-specific siRNA molecules in systemic leaves of wheat plant. Subsequent Pt inoculation resulted in a suppressed disease phenotype and a reduction in endogenous transcript levels of the targeted fungal genes indicating translocation of siRNA molecules from host to fungal cells. Efficiency of this host-generated trans-specific RNAi was enhanced by using BSMV silencing vectors defective in coat protein coupled with introducing fungal gene sequences simultaneously in sense and antisense orientation. The disease suppression indicated the likely involvement of these fungal genes in pathogenicity. This study demonstrates that BSMV-mediated in planta-generated RNAi is an effective strategy for functional genomics in rust fungi.

Journal

Plant Molecular BiologySpringer Journals

Published: Feb 17, 2013

References

  • Virus-induced gene silencing (VIGS) of genes expressed in root, leaf, and meiotic tissues of wheat
    Bennypaul, HS; Mutti, JS; Rustgi, S; Kumar, N; Okubara, PA; Gill, KS
  • Wheat leaf rust caused by Puccinia triticina
    Bolton, MD; Kolmer, JA; Garvin, DF
  • Leaf rust resistance gene Lr1, isolated from bread wheat (Triticum aestivum L.) is a member of the large psr567 gene family
    Cloutier, S; McCallum, B; Loutre, C; Banks, T; Wicker, T; Feuillet, C; Keller, B; Jordan, M
  • Virus-induced gene silencing-based functional characterization of genes associated with powdery mildew resistance in barley
    Hein, I; Barciszewska-Pacak, M; Hrubikova, K; Williamson, S; Dinesen, M; Soenderby, IE; Sundar, S; Jarmolowski, A; Shirasu, K; Lacomme, C
  • Barley stripe mosaic virus-induced gene silencing in a monocot plant
    Holzberg, S; Brosio, P; Gross, C; Pogue, GP
  • Generation of a wheat leaf rust, Puccinia triticina, EST database from stage-specific cDNA libraries
    Hu, G; Linning, R; McCallum, B; Banks, T; Cloutier, S; Butterfield, Y; Liu, J; Kirkpatrick, R; Stott, J; Yang, G; Smailus, D; Jones, S; Marra, M; Schein, J; Bakkeren, G
  • Hordeivirus replication, movement, and pathogenesis
    Jackson, AO; Lim, H-S; Bragg, J; Ganesan, U; Lee, MY

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