Infection-Associated Nuclear Degeneration in the Rice Blast Fungus Magnaporthe oryzae Requires Non-Selective Macro-Autophagy

Abstract

BackgroundThe rice blast fungus Magnaporthe oryzae elaborates a specialized infection structure called an appressorium to breach the rice leaf surface and gain access to plant tissue. Appressorium development is controlled by cell cycle progression, and a single round of nuclear division occurs prior to appressorium formation. Mitosis is always followed by programmed cell death of the spore from which the appressorium develops. Nuclear degeneration in the spore is known to be essential for plant infection, but the precise mechanism by which it occurs is not known.Methodology/Principal FindingsIn yeast, nuclear breakdown requires a specific form of autophagy, known as piecemeal microautophagy of the nucleus (PMN), and we therefore investigated whether this process occurs in the rice blast fungus. Here, we report that M. oryzae possesses two conserved components of a putative PMN pathway, MoVac8 and MoTsc13, but that both are dispensable for nuclear breakdown during plant infection. MoVAC8 encodes a vacuolar membrane protein and MoTSC13 a peri-nuclear and peripheral ER protein.Conclusions/SignificanceWe show that MoVAC8 is necessary for caffeine resistance, but dispensable for pathogenicity of M. oryzae, while MoTSC13 is involved in cell wall stress responses and is an important virulence determinant. By functional analysis of ΔMoatg1 and ΔMoatg4 mutants, we demonstrate that infection-associated nuclear degeneration in M. oryzae instead occurs by non-selective macroautophagy, which is necessary for rice blast disease.

Nucleophagy at a glance

“…In contrast to PMN, nuclear degradation in M. oryzae is dependent on core autophagy genes, such as M. oryzae ATG1 or ATG4 (Kershaw and Talbot, 2009). In contrast to large ring-like autophagosomes that mediate the degradation of A. oryzae nuclei, only smaller, more punctate autophagosomes surround the target nucleus during appressorium formation in M. oryzae (He et al, 2012). Moreover, the nucleus does not appear to be fully degraded, suggesting that nuclear breakdown might be performed by distinct macroautophagy-dependent processes in filamentous fungi (He et al, 2012).…”

“…To be able to infect rice plants, M. oryzae forms a specialized structure, the appressorium, which generates immense intracellular turgor pressure that results in the direct rupture of the plant cuticle and, thereby, facilitates fungal entry and colonization of plant tissue . Appressorium development involves a number of steps, one of which is nuclear degeneration in the spore from which the appressorium develops (see Poster) (Veneault-Fourrey et al, 2006;He et al, 2012). Although, M. oryzae possesses homologs of Vac8 and Tsc13, components of a putative PMN pathway, an Nvj1 homolog has not been identified and there is no evidence for the formation of NV junctions and for PMN activity.…”

“…Although, M. oryzae possesses homologs of Vac8 and Tsc13, components of a putative PMN pathway, an Nvj1 homolog has not been identified and there is no evidence for the formation of NV junctions and for PMN activity. Moreover, both M. oryzae Vac8 and Tsc13 are dispensable for nuclear breakdown during plant infection (He et al, 2012). In contrast to PMN, nuclear degradation in M. oryzae is dependent on core autophagy genes, such as M. oryzae ATG1 or ATG4 (Kershaw and Talbot, 2009).…”

“…Studies in both yeast and mammalian cells support the contention that selective digestion of portions of the nucleus can occur in order to maintain nuclear integrity (Roberts et al, 2003;Park et al, 2009;Mijaljica et al, 2012). By contrast, in other systems, such as (See poster insert) the ciliated protozoan Tetrahymena thermophila and some filamentous fungi, the evidence suggests that selective digestion of whole nuclei occurs as part of a program of cellular development or mechanism of infection (Akematsu et al, 2010;Shoji et al, 2010;He et al, 2012). Autophagy is a homeostatic intracellular degradation system widely conserved in eukaryotes that is known to operate in a selective manner to remove organelles, such as mitochondria or peroxisomes, and which, as will be discussed below, can accomplish the degradation of nucleusderived cargo or whole nuclei.…”

Carbon and nitrogen depletion-induced nucleophagy and selective autophagic sequestration of a whole nucleus in multinucleate cells of the filamentous fungus <i>Aspergillus oryzae</i>

“…Additionally, it was reported that nucleophagy of a whole nucleus occurs during appressorium formation required for plant infection in the rice blast fungus Magnaporthe oryzae (He et al, 2012) and after vegetative cell fusion and during nitrogen starvation in another plant pathogen Fusarium oxysporum (Corral-Ramos et al, 2015). In A. oryzae, the morphological process of nucleophagy has been characterized by fluorescence microscopy under the expression of an enhanced green fluorescence protein (EGFP)-fused AoAtg8, an orthologue of the autophagsomal marker S. cerevisiae Atg8, (EGFP-AoAtg8) and the nuclear marker mDsRed-fused histone H2B (H2B-mDsRed) (Shoji et al, 2010); AoAtg8-positive crescent-and cup-shaped autophagosome precursors are formed in the vicinity of the mDsRed-labeled nucleus, and then a ring-like autophagosome engulfs the whole nucleus.…”