Nitrogen-to-phosphorus (N:P) ratios are critical factors that influence harmful cyanobacterial blooms, but their roles in determining bloom magnitude, frequency, and duration require further study. In N-limited environments, N-fixing cyanobacteria, such as Aphanizomenon flos-aquae, can outcompete other phytoplankton. However, their ability to produce toxins appears to diminish when they fix nitrogen. The degree of N deficiency experienced by A. flos-aquae likely dictates the extent of N fixation and may also influence toxin production, and these dynamics may be further modulated by light availability. To investigate these interactions, we conducted a fully factorial experiment examining the responses of A. flos-aquae to selective pressures under strictly N-limited conditions. Batch cultures were grown under varying N:P ratios (5, 10, and 20) and light intensities (20, 50, 100 µmol-2s-1). Each treatment combination was replicated five times, and we measured chlorophyll-a (Chl-a), particulate carbon, particulate nitrogen ratios, particulate phosphorus (PP), and cylindrospermopsin production. We hypothesized that the highest N:P treatments grown in the highest light would yield the greatest biomass and cylindrospermopsin production rates despite having the highest C:N ratios. Similarly, we hypothesized that the lowest N:P grown at the lowest light would have the lowest C:N ratios but produce the least biomass and cylindrospermopsin. Our experiment was designed to determine whether light intensity and nutrient availability exert additive, interactive, or no effects on A. flos-aquae N fixation and toxin production by characterizing its physiological demands under N-limiting conditions.