Abstract
SummaryPost‐translational farnesylation can regulate subcellular localization and protein–protein interaction in eukaryotes. The function of farnesylation is not well identified in plant pathogenic fungi, particularly during the process of fungal infection. Here, through functional analyses of the farnesyltransferase β‐subunit gene, RAM1, we examine the importance of protein farnesylation in the rice blast fungus Magnaporthe oryzae. Targeted disruption of RAM1 resulted in the reduction of hyphal growth and sporulation, and an increase in the sensitivity to various stresses. Importantly, loss of RAM1 also led to the attenuation of virulence on the plant host, characterized by decreased appressorium formation and invasive growth. Interestingly, the defect in appressoria formation of the Δram1 mutant can be recovered by adding exogenous cAMP and IBMX, suggesting that RAM1 functions upstream of the cAMP signalling pathway. We found that two Ras GTPases, RAS1 and RAS2, can interact with Ram1, and their plasma membrane localization was regulated by Ram1 through their C‐terminal farnesylation sites. Adding a farnesyltransferase inhibitor Tipifarnib can result in similar defects as in Δram1 mutant, including decreased appressorium formation and invasive growth, as well as mislocalized RAS proteins. Our findings indicate that protein farnesylation regulates the RAS protein‐mediated signaling pathways required for appressorium formation and host infection, and suggest that abolishing farnesyltransferase could be an effective strategy for disease control.
Highlights
Protein farnesylation is one type of prenylation modification, which is required for the proper localization of many proteins in signal transduction, including Ras proteins (Maurer-Stroh et al, 2003; Zhang and Casey, 1996)
Deletion of the FTase α-subunit is lethal in fungi such as Saccharomyces cerevisiae and Candida albicans (He et al, 1991; Song and White, 2003), functions of farnesylation have been investigated by successful deletion of the FTase β-subunit in several fungi, including S. cerevisiae, C. albicans, Schizosaccharomyces pombe, Cryptococcus neoformans and Aspergillus fumigatus (He et al, 1991; Norton et al, 2017; Vallim et al, 2004; Yang et al, 2000)
The results showed that MoRam1 protein shares a 61% amino acid identity to that of Colletotrichum graminicola, 61% to F. oxysporum, 51% to A. fumigatus, 33% to C. neoformans, 34% to S. cerevisiae, 31% to C. albicans, 42% to Caenorhabditis elegans and 41% to Homo sapiens (Fig. S2, see Supporting Information) at the protein level with more than 65% query coverage
Summary
Protein farnesylation is one type of prenylation modification, which is required for the proper localization of many proteins in signal transduction, including Ras proteins (Maurer-Stroh et al, 2003; Zhang and Casey, 1996). The FTase is an αβ heterodimer composed of an essential α-subunit Ram and a non-essential β-subunit Ram (Casey and Seabra, 1996; Maurer-Stroh et al, 2003). Deletion of the FTase α-subunit is lethal in fungi such as Saccharomyces cerevisiae and Candida albicans (He et al, 1991; Song and White, 2003), functions of farnesylation have been investigated by successful deletion of the FTase β-subunit in several fungi, including S. cerevisiae, C. albicans, Schizosaccharomyces pombe, Cryptococcus neoformans and Aspergillus fumigatus (He et al, 1991; Norton et al, 2017; Vallim et al, 2004; Yang et al, 2000). The functions of farnesylation, especially during the infection process, are still largely unknown in plant pathogenic fungi
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