title: Leucoagaricus gongylophorus uses leaf-cutting ants to vector proteolytic enzymes towards new plant substrate.
abstract: The mutualism between leaf-cutting ants and their fungal symbionts revolves around processing and inoculation of fresh leaf pulp in underground fungus gardens, mediated by ant fecal fluid deposited on the newly added plant substrate. As herbivorous feeding often implies that growth is nitrogen limited, we cloned and sequenced six fungal proteases found in the fecal fluid of the leaf-cutting ant Acromyrmex echinatior and identified them as two metalloendoproteases, two serine proteases and two aspartic proteases. The metalloendoproteases and serine proteases showed significant activity in fecal fluid at pH values of 5-7, but the aspartic proteases were inactive across a pH range of 3-10. Protease activity disappeared when the ants were kept on a sugar water diet without fungus. Relative to normal mycelium, both metalloendoproteases, both serine proteases and one aspartic protease were upregulated in the gongylidia, specialized hyphal tips whose only known function is to provide food to the ants. These combined results indicate that the enzymes are derived from the ingested fungal tissues. We infer that the five proteases are likely to accelerate protein extraction from plant cells in the leaf pulp that the ants add to the fungus garden, but regulatory functions such as activation of proenzymes are also possible, particularly for the aspartic proteases that were present but without showing activity. The proteases had high sequence similarities to proteolytic enzymes of phytopathogenic fungi, consistent with previous indications of convergent evolution of decomposition enzymes in attine ant fungal symbionts and phytopathogenic fungi.
authors.affiliation: Centre for Social Evolution, Department of Biology, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Pepijn W
authors.initials: PW
authors.lastname: Kooij
authors.affiliation: Protein Research Group, Department of Biochemistry and Molecular Biology, University of Southern Denmark, Odense, Denmark.
authors.forename: Adelina
authors.initials: A
authors.lastname: Rogowska-Wrzesinska
authors.affiliation: Centre for Social Evolution, Department of Biology, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Daniel
authors.initials: D
authors.lastname: Hoffmann
authors.affiliation: Protein Research Group, Department of Biochemistry and Molecular Biology, University of Southern Denmark, Odense, Denmark.
authors.forename: Peter
authors.initials: P
authors.lastname: Roepstorff
authors.affiliation: Centre for Social Evolution, Department of Biology, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Jacobus J
authors.initials: JJ
authors.lastname: Boomsma
authors.affiliation: Centre for Social Evolution, Department of Biology, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Morten
authors.initials: M
authors.lastname: Schiøtt
mesh_terms: D000363:Agaricales / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D000818:Animals; D001000:Ants / Q000382:microbiology* / Q000502:physiology; D005075:Biological Evolution; D010447:Peptide Hydrolases / Q000235:genetics / Q000378:metabolism; D018515:Plant Leaves; D013559:Symbiosis
chemical_list: D010447:Peptide Hydrolases