Fungi are ubiquitous in terrestrial ecosystems and play an important role in biogeochemical cycling because of their function as litter decomposers. It has been demonstrated that increased nitrogen (N) deposition decreases fungal biomass and changes the relative abundance of particular groups and species. In addition increased N can slow litter decomposition and reduce lignolytic enzyme activity. To understand the functioning of fungi under increased N addition in more detail, we performed a soil metatranscriptomic analysis on forest floor samples from a chronic N-addition experiment at Harvard Forest (MA, USA). We extracted total RNA, followed by reverse transcription of poly-A tailed mRNA and Illumina paired-end sequencing of the cDNA.
Results/Conclusions
In the forest floor of control and nitrogen amended plots, a total of ~460,000 genes were expressed at the time of sampling, of which 172,000 and 95,000 were uniquely expressed in the nitrogen and control plots, respectively. A total of ~23,000 genes (5% of all expressed genes) were either significantly up-regulated or down-regulated in the control versus the N-amended plots (P<0.05). Of these 23,000 genes, we focused on genes with a medium to high expression value (RPKM >5, Reads Per Kilobase of exon per Million mapped sequence reads) in either the control or N-amended plots, so that there would be at least a 2-fold change in expression. Most of the differences in expression were found within primary metabolic and cellular processes. More specifically, several genes closely related to fungal glycoside hydrolases, laccases and peroxidases, involved in lignocellulose degradation, were found to be down-regulated in the N-amended plots compared to the control plots. Within our study, metatranscriptomics proved to be a useful technique to study the activity of specific genes of interest, while at the same time revealing shifts in the activity of genes that would be overlooked in gene-specific qPCR analysis.