Proteolytic Enzyme Activity in Temperate Forest Soils at Harvard Forest and Pisgah State Forest 2007-2010

The objective of this research is to investigate the processes that limit or promote the activity and production of proteolytic enzymes in temperate forest soils. To meet this objective, we performed a series of integrated observations and experiments to investigate a conceptual model of proteolytic enzyme activity whereby activity is a function of the interaction between four parameters: soil temperature and moisture, substrate concentration, and the enzyme pool size. We used four dominant temperate forest tree species that differ in SOM chemistry and the enzymatic capabilities of their fungal symbionts as a model system. These four species differed in mycorrhizal association, with white ash (Fraxinus americana) and sugar maple (Acer saccharum) supporting arbuscular mycorrhizal (AM) fungi and eastern hemlock (Tsuga canadensis) and American beech (Fagus grandifolia) supporting ectomycorrhizal (ECM) fungi. Further, the ECM associated species have leaf litter and SOM that is characterized by higher ratios of C:N than the AM associated. Soil samples were collected two sites, one located at the Prospect Hill Tract of the Harvard Forest and the other at the Pisgah State Forest. The sites have similar land use history and stand age. Soils at both sites are inceptisols classified as Typic Dystrochrepts derived from glacial till overlying granite-schist-gneiss bedrock. Experimental plots dominated by one of four target tree species were established at each site. Stands of sugar maple (Acer saccharum) and American beech (Fagus grandifolia) were located in Pisgah. Stands of Eastern hemlock (Tsuga canadensis) and white ash (Fraxinus americana) were located in Harvard Forest. At a later date, plots were also established in a red oak (Quercus rubra) stand on Prospect Hill in the MES tower footprint. At each site we located six replicate, 8 m radius, monodominant plots that were based on the following criteria: (1) more than 80% of the standing basal area was composed of the target tree species, (2) the fresh litterfall layer was dominated by the target species, and (3) the core 5 m of the plot contained only the target tree species. Significant organic horizons were present only in the beech and hemlock plots. The lack of organic horizon in the white ash plots may be due to the presence of earthworms, which are not present in the other plots. In 2008, we investigated how temperature and substrate availability impact the activity of proteolytic enzymes in temperate forest soils, and whether the activity of proteolytic enzymes and other enzymes involved in the acquisition of N (i.e., chitinolytic and ligninolytic enzymes) differs between trees species that form associations with either ECM or AM fungi. We performed a factorial lab experiment using soils sampled at three time points over the growing season, where we assayed proteolytic rates at two temperatures (field temperature at time of sampling and 23 deg C) and two substrate levels (ambient or elevated protein in the form of casein). Further, we performed an in-growth experiment in the field to investigate differences between ECM and AM tree roots in their effects on enzyme production and activity. In 2009, we assayed the temperature sensitivity of proteolytic enzymes at three time points over the growing season. We quantified proteolytic enzyme activity at six temperatures ranging from 0-35 deg C in the soils from all four species. Finally, in 2010, we assayed the activity of proteolytic, chitinolytic, and ligninolytic enzymes in the rhizosphere and bulk soil of all four tree species. We found that proteolytic activity was more limited by substrate than by temperature, with declines in both limitations as soil temperature increased over the growing season. Fine roots stimulated proteolytic activity in the rhizosphere, the zone immediately around the root, likely by enhancing microbial enzyme production. Ectomycorrhizal roots stimulated activity more than arbuscular mycorrhizal roots. The results of this research suggest that climate warming in the absence of increases in substrate availability will have a modest effect on proteolytic activity in temperate forests. Further, global changes that alter belowground carbon allocation by trees are likely to have a larger impact on N cycling in ectomycorrhizal stands than in arbuscular mycorrhizal stands.