Plants often experience low oxygen conditions as the consequence of reduced oxygen availability in their environment or due to a high activity of respiratory metabolism. Recently, an oxygen sensing pathway was described in Arabidopsis thaliana which involves the migration of an ERF transcription factor (RAP2.12) from the plasma membrane to the nucleus upon hypoxia. Moreover, RAP2.12 protein level is controlled through an oxygen-dependent branch of the N-end rule pathway for proteasomal degradation. Inside the nucleus, RAP2.12 induces the expression of genes involved in the adaptation to reduced oxygen availability. In the present study, we describe the oxygen concentration and time-resolved characterization of RAP2.12 activity. A reduction of the oxygen availability to half the concentration in normal air is sufficient to trigger RAP2.12 relocalization into the nucleus, while nuclear accumulation correlates with the first induction of the molecular response to hypoxia. Nuclear presence of RAP2.12 may not only depend on relocalization of existing protein, but involves de novo synthesis of the transcription factor as well. After re-oxygenation of the tissue, degradation of RAP2.12 in the nucleus was observed within 3 h, concomitant with reduction in hypoxia responsive gene transcripts to normoxic levels.