Centre for Social Evolution, Department of Biology, University of Copenhagen, Copenhagen, Denmark.
Protein Research Group, Department of Biochemistry and Molecular Biology, University of Southern Denmark, Odense, Denmark.
ISME J. 2014 May;8(5):1032-40. doi: 10.1038/ismej.2013.231. Epub 2014 Jan 9.
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.
切叶蚁与其真菌共生体之间的互利共生关系围绕着在地下真菌园中处理和接种新鲜叶浆展开，这一过程由蚂蚁粪便中的液体介导，这些液体被沉积在新添加的植物基质上。由于食草性进食通常意味着生长受到氮的限制，我们克隆并测序了在切叶蚁 Acromyrmex echinatior 的粪便中发现的六种真菌蛋白酶，并将它们鉴定为两种金属内肽酶、两种丝氨酸蛋白酶和两种天冬氨酸蛋白酶。金属内肽酶和丝氨酸蛋白酶在 pH 值为 5-7 的粪便中表现出显著的活性，但天冬氨酸蛋白酶在 pH 值为 3-10 的范围内没有活性。当蚂蚁被限制在不含真菌的糖水饮食中时，蛋白酶活性消失。与正常菌丝体相比，两种金属内肽酶、两种丝氨酸蛋白酶和一种天冬氨酸蛋白酶在专门的菌丝尖端——菌褶中上调，菌褶是菌丝体的特化部分，其唯一已知的功能是为蚂蚁提供食物。这些综合结果表明，这些酶来源于被蚂蚁摄入的真菌组织。我们推断，这五种蛋白酶可能有助于加速从蚂蚁添加到真菌园的叶浆中的植物细胞中提取蛋白质，但也可能具有调节功能，例如激活酶原，尤其是那些存在但没有表现出活性的天冬氨酸蛋白酶。这些蛋白酶与植物病原菌真菌的蛋白水解酶具有高度的序列相似性，这与分解酶在拟巢菌目蚂蚁和植物病原菌真菌中的趋同进化的先前指示一致。
