The ER Chaperone LHS1 Is Involved in Asexual Development and Rice Infection by the Blast FungusMagnaporthe oryzae     

Abstract

In planta secretion of fungal pathogen proteins, including effectors destined for the plant cell cytoplasm, is critical for disease progression. However, little is known about the endoplasmic reticulum (ER) secretion mechanisms used by these pathogens. To determine if normal ER function is crucial for fungal pathogenicity, Magnaporthe oryzae genes encoding proteins homologous to yeast Lhs1p and Kar2p, members of the heat shock protein 70 family in Saccharomyces cerevisiae, were cloned and characterized. Like their yeast counterparts, both LHS1 and KAR2 proteins localized in the ER and functioned in an unfolded protein response (UPR) similar to the yeast UPR. Mutants produced by disruption of LHS1 were viable but showed a defect in the translocation of proteins across the ER membrane and reduced activities of extracellular enzymes. The Dlhs1 mutant was severely impaired not only in conidiation, but also in both penetration and biotrophic invasion in susceptible rice (Oryza sativa) plants. This mutant also had defects in the induction of the Pi-ta resistance gene-mediated hypersensitive response and in the accumulation of fluorescently-labeled secreted effector proteins in biotrophic interfacial complexes. Our results suggest that proper processing of secreted proteins, including effectors, by chaperones in the ER is requisite for successful disease development and for determining host-pathogen compatibility via the gene-for-gene interaction.

Translocation ofMagnaporthe oryzaeEffectors into Rice Cells and Their Subsequent Cell-to-Cell Movement

“…The importance of effector accumulation in BICs to the compatible interaction is highlighted by the general absence of fluorescent effectors in BICs in the incompatible interaction (Mosquera et al, 2009;Yi et al, 2009). Although faint fluorescence from effector:FPs was sometimes observed in cells undergoing R gene-mediated HR, fully developed fluorescent BICs were not observed.…”

“…However, we cannot rule out a role for differential effector expression because it remains possible that proteins that are highly expressed in BIC-associated cells are redistributed throughout the IH by cytoplasmic streaming through septal pores. At least initially, AVR-Pita1 appears to be secreted into BICs using the normal fungal ER-mediated secretion machinery because mutation in the ER chaperone LHS1 with a role in secretion severely impairs its BIC accumulation and its function in triggering Pita-mediated HR (Yi et al, 2009). Another study suggested that M. oryzae uses different secretion mechanisms in planta because the APT2 gene, which encodes a Golgi-localized P-type ATPase, appears to only be involved in secretion of the tested AVR effector and a subset of extracellular enzymes (Gilbert et al, 2006).…”

“…Recently, Yi et al (2009) have shown that M. oryzae mutants lacking the lumenal heat shock protein seventy (LHS1) gene, which encodes an endoplasmic reticulum (ER) chaperone functioning in protein secretion, were severely impaired in biotrophic invasion and in induction of R genemediated HR. These same mutants showed minor impairment of axenic growth in nutrient medium.…”

Nonself Recognition Through Intermolecular Disulfide Bond Formation of Ribonucleotide Reductase in Neurospora

“…The presence of the hygunPA(788-923)-HA protein in the pellet fraction (although less than observed in the self-incompatible colony) from the self-compatible colony may be due to our use of a trpC promoter to drive fusion protein expression. Previous studies have observed that expression from strong, constitutive promoters causes some overexpressed protein to become insoluble (Yi et al 2009;Tegel et al 2011), including studies that examine fungal protein complexes involved in nonself recognition (Mathur et al 2012).…”

- Enhanced Disease Susceptibility 1 and Salicylic Acid Act Redundantly to Regulate Resistance Gene-Mediated Signaling

“…Total RNA was isolated from the frozen fungal and plant tissues with Easy-spin TM total RNA extraction kit (iNtRON Biotechnology, Seoul, Korea) according to the manufacturer's instruction. To quantify levels of transcript, quantitative RT-PCR was performed as described [78]. Briefly, 5 mg of total RNA was reverse transcribed into first-strand cDNA with oligo (dT) primer using SuperScript TM First-Strand Synthesis System (Invitrogen TM Life Technologies, Carlsbad, CA, USA) according to the manufacturer's instruction.…”