Wheat plants (Triticum aestivum L.) were subjected to varying phosphorus (P) supply and canopy ¹³CO₂ feeding to uncouple the plant and soil factors regulating carbon (C) allocation between the shoot and roots and in the P-enriched v. P-deficient soil zone. In a split-root system, transferring from 200/200 µM P (high/high) to high/nil P or nil/nil P for 7 days was associated with 18–30% increase in the ratio of root-to-total ¹³C, whereas 8–12% more ¹³C was retained in the shoot of plants grown under continuous high/high P. Although the C signal between the shoot and roots weakened at day 12, it closely correlated with root P acquisition at both days. In a non-split-root system, plants supplied with 200 μm P (high) for 7 and 12 days had a lower ratio of root-to-total ¹³C than plants with continuous 20 μm P (low). Preferential C allocation and increased P acquisition occurred before any measurable growth difference. Shoot P status had a greater influence than soil P supply on plant C allocation, and rapid C signalling between the shoot and roots might serve as an important component of plant response to heterogeneous P conditions.