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Protease inhibition by Heterodera glycines cyst content: evidence for effects on the Meloidogyne incognita proteasome

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Proteases from Heterodera glycines and Meloidogyne incognita juveniles were inhibited by heat-stable content from H. glycines cysts (hHglCE), and by a polyphenol (EGCG) similar to a compound previously identified in Globodera cysts. General protease activities detected using the nematode peptide KSAYMRFa were inhibited by EGCG (IC50 1.19 mM, H. glycines; 0.34 mM, M. incognita) but not by hHglCE. However, hHglCE and EGCG each inhibited proteasome-associated chymotrypsin-like (CT-L) activity. EGCG IC50 values were 0.47 mM (H. glycines) and 0.15 mM (M. incognita). hHglCE IC50 values were 0.16 and 0.005 mM hHglCEeq μl−1 for H. glycines and M. incognita, respectively. Across all substrate-inhibitor combinations, M. incognita proteases were inhibited more robustly than those from H. glycines, particularly by hHglCE. In addition to CT-L protease, post-glutamate peptide hydrolysing (PGPH) and trypsin-like (T-L) proteasome proteases were detected in M. incognita, and each of these was also strongly inhibited by hHglCE. hHglCE inhibited CT-L, PGPH and T-L proteases within catalytic subunits from yeast (Saccharomyces cerevisiae) and human proteasomes. Proteasome inhibitors MG-132 and aclacinomycin A each inhibited M. incognita CT-L and PGPH activities by more than 80% at 20-100 μM, and hHglCE inhibited the same proteases by 70-80% at 0.04 hHglCEeq μl−1. hHglCE completely inhibited M. incognita T-L activity, but CT-L activity in native content from H. glycines cysts was not inhibited. Evidence that H. glycines cysts contain inhibitors of all proteases associated with the proteasome establishes the cyst as an important new target to explore for potential nematode control compounds. In addition, characterisation of protease activities from a core cellular metabolic component using M. incognita is novel for plant-parasitic nematodes.

Affiliations: 1: USDA-ARS Nematology Laboratory, Beltsville, MD, USA

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2015-01-12
2017-11-23

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