Nitrogen (N) serves both as a vital macronutrient and a signaling molecule for plants. Unveiling key regulators involved in N metabolism helps dissect the mechanisms underlying N metabolism, which is essential for developing maize with high N use efficiency. Two maize lines, B73 and Ki11, show differential chlorate and low-N tolerance. Time-course transcriptomic analysis reveals that the expression of N utilization genes (NUGs) in B73 and Ki11 have distinct responsive patterns to nitrate variation. By the coexpression networks, significant differences in the number of N response modules and regulatory networks of transcription factors (TFs) are revealed between B73 and Ki11. There are 23 unique TFs in B73 and 41 unique TFs in Ki11. MADS26 is a unique TF in the B73 N response network, with different expression levels and N response patterns in B73 and Ki11. Overexpression of MADS26 enhances the sensitivity to chlorate and the utilization of nitrate in maize, at least partially explaining the differential chlorate tolerance and low-N sensitivity between B73 and Ki11. The findings in this work provide unique insights and promising candidates for maize breeding to reduce unnecessary N overuse.