Anecdotal observations of improved glyphosate efficacy on glyphosate-resistant (GR) tall waterhemp populations in corn production compared with soybean suggested the presence of nitrogen (N) fertilizer may influence the expression of glyphosate resistance. Greenhouse and field experiments were conducted to determine the influence of soil-applied nitrogen fertilizer on the growth rate and sensitivity of glyphosate-susceptible (GS) and GR tall waterhemp and Palmer amaranth. The addition of supplemental fertilizer increased the relative growth rate (plant height and shoot volume), number of nodes, and percentage of shoot nodes with axillary branches on GS and GR biotypes of both weed species. The axillary bud activity was increased 52 and 8% with increasing N for the GR and GS biotypes of tall waterhemp and Palmer amaranth, respectively. The GS populations of tall waterhemp and Palmer amaranth were more sensitive to glyphosate in the greenhouse under increased fertilizer levels compared with no fertilizer. Additionally, GR tall waterhemp was more sensitive to glyphosate under the higher fertilizer treatments, which resulted in a reduction in the calculated resistance factor (RF) from 27.8 under no fertilizer to 4.7 for the high fertilizer treatment. The RF for GR Palmer amaranth was not influenced by the fertilizer treatments in the greenhouse. Field experiments demonstrated that glyphosate efficacy may be greater on GR populations of tall waterhemp and Palmer amaranth under high N conditions, but these results were not consistent and most likely were influenced by soil moisture in 2012, which was more limiting than N supply. This research implies that soil fertility can influence the sensitivity of some GR weed species to glyphosate and the RF. Therefore, the evolution and management of GR weed species in commercial crop production may be influenced by the nutrient status of the soil and the use of supplemental fertilizers.