Figure 6.

Leaf nitrogen remobilization is interrupted during vegetative stage in the Osatg7-1 mutant. Nitrogen flow in the 10th leaf of OsATG7+/+ and Osatg7-1 was estimated by ¹⁵N pulse chase analysis using elemental analyzer/isotope ratio mass spectrometry. Total nitrogen and ¹⁵N atom% excess of the 10th leaves were measured at three time points (just after leaf emergence, full expansion, and senescence; A). ¹⁵N-labeled (¹⁵NH₄)₂SO₄ was supplied for 3 d (indicated as gray slashed column) when the 10th leaves were emerged. After ¹⁵N supply, culture medium was changed to pH-adjusted water until the 10th leaves were fully expanded (for 3 d in OsATG7+/+ and for 4 d in Osatg7-1). From full expansion onward, nutrient solution including nonlabeled nitrogen was supplied once a week until 30 d after 10th leaf emergence. White and black symbols, Total nitrogen amount and ¹⁵N atom% excess, on left and right axes, respectively; circles, OsATG7+/+; squares, Osatg7-1. The origin of nitrogen flow in the 10th leaves during leaf expansion (a) was estimated in B. Nitrogen influx and efflux during leaf senescence (b) were also estimated in C. In each graph, data are means ± se (n = 3–4). Asterisks indicate significant difference analyzed by Student’s t test (P < 0.05). Calculations in B and C are described in “Materials and Methods.” n.s., Not significant.