Generation of reactive oxygen species by fungal NADPH oxidases is required for rice blast disease

Abstract

One of the first responses of plants to microbial attack is the production of extracellular superoxide surrounding infection sites. Here, we report that Magnaporthe grisea, the causal agent of rice blast disease, undergoes an oxidative burst of its own during plant infection, which is associated with its development of specialized infection structures called appressoria. Scavenging of these oxygen radicals significantly delayed the development of appressoria and altered their morphology. We targeted two superoxide-generating NADPH oxidaseencoding genes, Nox1 and Nox2, and demonstrated genetically, that each is independently required for pathogenicity of M. grisea. ⌬nox1 and ⌬nox2 mutants are incapable of causing plant disease because of an inability to bring about appressorium-mediated cuticle penetration. The initiation of rice blast disease therefore requires production of superoxide by the invading pathogen.O ne of the earliest manifestations of the plant defense response is the production of reactive oxygen species (ROS), including superoxide and its dismutation product, hydrogen peroxide (1) . These ROS can kill pathogens directly (2) but also strengthen plant cell walls through the oxidative cross-linking of cell wall structural proteins (3) and may function in the regulation of programmed cell death (4). Although numerous studies have documented the detection of plant-derived ROS (2, 3, 5-7), very little is known about the role of ROS generation in invading plant pathogenic microorganisms. However, the recent discovery of functional members of the superoxide-generating NADPH oxidase (Nox) family within filamentous fungi (8) has led to increased speculation regarding the possible role of ROS in pathogenic species.The most well characterized Nox remains that of the human phagocytic leukocyte, a multisubunit oxidase formed by the cytosolic regulatory components Rac, p67 phox , p47 phox , and p40 phox and the integral membrane protein flavocytochrome b 558 , composed of the catalytic subunit gp91 phox and p22 phox (9). In activated macrophages, Nox enzymes induce K ϩ influx, causing pH changes in the phagocytic vacuole, leading to the killing of pathogens through activation of neutral proteases (10). Mutations in the catalytic gp91 phox subunit result in chronic granulomatous disease, an immunological disorder in which macrophages are unable to prevent the spread of infection (11). Plants contain enzymes that are homologous to gp91 phox , designated respiratory burst oxidase homologues (Rboh) (12). Arabidopsis thaliana, for example, possesses 10 Rboh isoforms involved in a diverse range of plant processes. ROS generated by the A. thaliana RHD2/RBOHC regulate root hair growth through the activation of Ca 2ϩ channels (13), whereas RBOHD and RBOHF regulate stomatal closure, seed germination, and root elongation through abscisic acid signaling (14). Recent studies have shown that it is the activation of RBOHD and RBOHF that is responsible for ROS accumulation in several plant-microbe interactions (15). However, r...

Unraveling the Tapestry of Networks Involving Reactive Oxygen Species in Plants

“…Three papers have brought to the fore the importance of ROS production and scavenging by the pathogen. Egan et al (2007) reported that the generation of ROS by the rice blast fungal NADPH oxidase is required for infection, whereas Molina and Kahmann (2007) documented that activation of ROS scavenging mechanisms by the fungal Ustilago maydis Yap1 gene is essential to overcome the plant defense mechanisms and allow infection. In addition, Mittapalli et al (2007) reported that the ROS scavenging mechanisms of the Hessian fly (Mayetiola destructor) could play an important function in the interaction with its wheat (Triticum aestivum) host.…”

Fungal Morphogenesis

“…It should be noted that there are examples of effectors with important roles in polarized hyphal growth that are not well conserved in S. cerevisiae. A particularly prominent example is NADPH oxidase, which appears to be an effector of Rac1 in filamentous fungi (Tanaka et al 2006;Semighini and Harris 2008), in which it regulates seemingly diverse morphogenetic processes, such as lateral branching and infection-related morphogenesis (Tanaka et al 2006;Egan et al 2007). Small GTPases, such as Ras, Rho, and Cdc42, do not operate in isolation, but rather in a sequential manner as suggested by studies in S. cerevisiae and S. pombe.…”

TheO-Mannosyltransferase PMT4 Is Essential for Normal Appressorium Formation and Penetration inUstilago maydis

“…To overcome this barrier, fungi have developed different mechanisms ranging from penetration through natural openings, such as stomata, to the formation of appressoria (Howard and Valent, 1996;Dean, 1997;Talbot, 2003), which are specialized structures that use mechanisms including osmotic pressure, hydrolytic enzymes, and the generation of reactive oxygen intermediates to break into host plant tissue (Howard et al, 1991;Egan et al, 2007;Skamnioti and Gurr, 2007).…”