Collaborating scientists at the U.S. Department of Agriculture’s Agricultural Research Service (USDA-ARS) and at Washington State University (WSU) have identified a protein that allows the fungus responsible for white mold stem rot in more than 600 plant species to overcome plant defenses.

The team’s study showed that the Sclerotinia sclerotiorum fungus produces the protein, SsPINE1, which directly inactivates polygalacturonase-inhibiting proteins (PGIPs) that plants use as their own natural defense against pathogens. The study findings could help researchers to develop a new, more precise system of control against S. sclerotiorum, which attacks potatoes, soybeans, sunflowers, peas, lentils, canola, and many other broad leaf crops. In a bad year, the damage caused by this fungus can add up to billions of dollars.

Commenting on the study results, Kiwamu Tanaka, PhD, an associate professor in Washington State University’s department of plant pathology, said, “I got goosebumps when we found this protein. It answered all these questions scientists have had for the last 50 years: Why these fungi always overcome plant defenses? Why do they have such a broad host range, and why are they so successful?”

Tanaka is co-author of the researchers’ published paper in Nature Communications, which is titled, “A fungal extracellular effector inactivates plant polygalacturonase-inhibiting protein.”

S. sclerotiorum fungi cause plants to rot and die by secreting chemicals called polygalacturonases (PG), which break down the plant’s cell walls. “Since plant cell wall is a major barrier to intrusion by pathogenic microorganisms, pathogens secret an array of cell wall-degrading enzymes (CWDEs) to compromise cell wall integrity and gain access,” the authors wrote. “An important part of the CWDEs is pectin-degrading PGs as pectin is an important component of cell wall and the middle lamella integrity in order to gain access.” In fact, they further commented, S. sclerotiorum has a genome that encodes at least five endopolygalacturonases that are expressed at different infection stages and conditions.

Plants have, in turn, evolved to protect themselves by producing PGIP, a protein that stops or inhibits PG produced by the pathogenic fungus. Since the discovery of PGIP in 1971 scientists have recognized that some fungal pathogens have a way to overcome a plant’s PGIP. But it’s not yet known exactly how. “To date, how fungal pathogens specifically overcome PGIP inhibition is unknown,” the investigators noted.

“What you have is essentially a continuous arms race between fungal pathogens and their plant hosts, an intense battle of attack, counterattack, and counter-counterattack in which each is constantly developing and shifting its chemical tactics in order to bypass or overcome the other’s defenses,” said research plant pathologist Weidong Chen, PhD, with the ARS Grain Legume Genetics Physiology Research Unit in Pullman, WA.

For their study, the researchers carried out transcriptome analysis and knockout screening of genes expressed preferentially at early stages of plant infection by both a wild type S. sclerotiorum strain, and a genetically defined oxalate-minus mutant. “The oxalate-minus mutant was included in the analysis because it retained pathogenicity despite previous claims that oxalic acid is the primary pathogenicity determinant,” the team explained.

Chen said the key to identifying the protein SsPINE1—Sclerotinia sclerotiorum PGIP-inactivating effector 1—was looking outside the cells. “We found it by looking at the materials excreted by the fungus,” he said. “And there it was. When we found this protein, SsPINE1, which interacted with PGIP, it made sense.”

Lab tests with Sclerotinia in which SsPINE1 had been deleted confirmed that the protein was responsible for permitting the fungus to bypass plants’ PGIP defenses. The experiments showed that SsPINE1 knockouts were dramatically less virulent than Sclerotinia that produced the protein.

The discovery of SsPINE1 has opened new avenues to investigate for controlling white mold stem rot pathogens, including the potential to develop more effective targeted breeding to make plants naturally resistant to sclerotinia diseases. The team’s reported research also confirmed that other related fungal pathogens use this counterstrategy, a finding that only serves to make the discovery even more important.

The “battle” between fungal PGs and plant PGIPs has been well documented since the discovery of PGIP, and has been thought to be “one-on-one engagement,” the scientists noted. The newly reported results introduce SsPINE1 as what they describe as “a third player … in the PG-PGIP battleground.” As the investigators further pointed out, current efforts in deploying PGIPs for improving plant resistance focus on boosting the expression of known PGIPs, as well as identifying or engineering novel PGIPs with enhanced potency and utility against a broader range of PGs. “Now with the knowledge of PGIP counter-inhibitors such as SsPINE1, newly engineered PGIPs that can avoid the recognition by SsPINE1 could be deployed for effective breeding of resistance plants against PINE1-possessing necrotrophic fungal pathogens,” they suggested.

The research is part of the National Sclerotinia Initiative, a multiorganization effort that ARS created to counterattack S. sclerotiorum because the fungus does so much damage around the world. The research team also included scientists from USDA-ARS, WSU, Northwestern A&F University in Shaanxi, China, Wuhan Polytechnic University, and Huazhong Agricultural University.

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