Phosphorus (P) is an essential macronutrient for plant growth. In deciduous trees, P is remobilized from senescing leaves and stored in perennial tissues during winter for further growth. Annual internal recycling and accumulation of P are considered an important strategy to support the vigorous growth of trees. However, the pathways of seasonal re‐translocation of P and the molecular mechanisms of this transport have not been clarified. Here we show the seasonal P re‐translocation route visualized using real‐time radioisotope imaging and the macro‐ and micro‐autoradiography. We analysed the seasonal re‐translocation P in poplar (Populus alba. L) cultivated under 'a shortened annual cycle system', which mimicked seasonal phenology in a laboratory. From growing to senescing season, sink tissues of 32P and/or 33P shifted from young leaves and the apex to the lower stem and roots. The radioisotope P re‐translocated from a leaf was stored in phloem and xylem parenchyma cells and redistributed to new shoots after dormancy. Seasonal expression profile of phosphate transporters (PHT1, PHT5 and PHO1 family) was obtained in the same system. Our results reveal the seasonal P re‐translocation routes at the organ and tissue levels and provide a foothold for elucidating its molecular mechanisms. Summary Statement: The phosphorus re‐translocation routes in Populus alba were visualized and gene expression of Pi transporters was analysed, using a shortened annual cycle system. Sink tissues of P shifted from young leaves to the stem and roots during senescence and stored P were reused for next spring growth. [ABSTRACT FROM AUTHOR]