Updated project metadata. Trees establish a symbiotic relationship with specialized soil fungi, called ectomycorrhizae, which is essential for nutrition, growth and health of temperate forest ecosystems. Understanding the mechanisms governing the establishment and functioning of ectomycorrhiza is important because of the role of forests in sequestering CO2 and also to develop ways to optimize tree productivity and sustainability. Here, we investigated the response of an oak species to ectomycorrhiza formation using a two dimensional differential in gel electrophoresis (2D-DIGE) and MALDI-TOF/TOF mass spectrometry proteomics approach. At the root level, changes in the abundance of 34 unique oak proteins were detected and revealed proteins involved in carbon and energy metabolism, protein processing and degradation, response to oxidative stress, lipid metabolism/transport, nitrogen and phosphorous assimilation and cell wall modification. Proteins supporting the importance of the secretory pathway functioning, in particular of the endoplasmic reticulum, during ectomycorrhiza functioning were identified. These proteins were identified as components of the endoplasmic reticulum folding/chaperoning machinery and proteins involved in the ER quality control system. This study constitutes an important contribution for the understanding of the mechanisms underlying the response of plants to ectomycorrhizal symbiosis establishment.