Gene expression and loss of function of GmbHLHm1 in soybean nodules. (A) Developmental expression (DAP, days after planting) of GmbHLHm1 in N2-fixing soybean nodules and nodule-detached roots. (B) GmbHLHm1 expression in leaves (L), nodules (N), and nodule-detached roots with (R +I) or without (R -I) rhizobia from 32-d-old plants grown without nitrogen. (C) Diel expression of GmbHLHm1 in nodules. (D and E) Cellular expression identified using a GmbHLHm1 promoter:GUS fusion in both infected and uninfected nodule cells (D) and a nodulated root (E). IR, infected region; P, parenchymal cells; VT, vascular trace. (F) Chlorotic shoots of plants grown solely on sat1-nodulated roots supplied with nutrient solution containing no nitrogen. (G) Reduction in leghemoglobin in infected cells of sat1 nodules. (H and I) Reduction in nodule number (n = 19; P = 0.0125) and nodule fresh weight (n = 11; P = 0.0013) in bhlhm1 transgenic events. Data represent means ± SE. *P < 0.05. (J and K) Toludine blue-stained fresh tissue cross-sections of vector and bHLHm1-transformed nodules highlighting small infected cells in bHLHm1 nodules. (L and M) TEM analysis of cross-sections of infected and uninfected cells from vector- and sat1-transformed nodules. IC, bacteroid-infected cell; UC, uninfected cell.