According to climate change scenarios, the seasonal course of temperature will change in most regions of the world, raising the question of how this will influence seasonal nitrogen (N) storage in deciduous trees. The key to this question is a detailed understanding of the underlying regulatory mechanisms, which was addressed in this study by analysing (i) the effects of low temperatures (13–1 °C) on bark storage protein (BSP) transcription, BSP and total protein accumulation and amino acid metabolism; (ii) the effects of interactions between low temperatures and photoperiod on these processes; and (iii) the regulatory role of amino acids in the bark. For this purpose, we exposed grey poplar trees (Populus × canescens) to three different treatments of changing photoperiod at constant temperature, changing temperature at constant photoperiod, and both changing photoperiod and temperature. Under a shortened photoperiod, a substantial increase of BSP transcripts was observed that was correlated with the accumulation of bark proteins, indicating a metabolic shift to promote long-term N storage. Irrespective of the applied photoperiod, exposure to low temperatures (5 or 1 °C) caused a strong increase of BSP transcripts, which was not paralled by significant increases of BSP and total bark proteins. We conclude that the interaction between effects of photoperiod and temperature is dependent on the carbon status of the trees, and reflects a metabolic adjustment of reduced carbon consumption for BSP synthesis. These results demonstrate the differential temperature sensitivity of processes involved in seasonal N storage, implying vulnerability to changing environmental conditions.