Invasive plants can increase soil nitrogen (N) pools and accelerate soil N cycling rates, but their effect on gross N cycling and nitrous oxide (N2 O) emissions has rarely been studied. We hypothesized that perennial pepperweed (Lepidium latifolium) invasion would increase rates of N cycling and gaseous N loss, thereby depleting ecosystem N and causing a negative feedback on invasion. We measured a suite of gross N cycling rates and net N2 O fluxes in invaded and uninvaded areas of an annual grassland in the Sacramento-San Joaquin River Delta region of northern California. During the growing season, pepperweed-invaded soils had lower microbial biomass N, gross N mineralization, dissimilatory nitrate reduction to ammonium (DNRA), and denitrification-derived net N2 O fluxes (P < 0.02 for all). During pepperweed dormancy, gross N mineralization, DNRA, and denitrification-derived net N2 O fluxes were stimulated in pepperweed-invaded plots, presumably by N-rich litter inputs and decreased competition between microbes and plants for N (P < 0.04 for all). Soil organic carbon and total N concentrations, which reflect pepperweed effects integrated over longer time scales, were lower in pepperweed-invaded soils (P < 0.001 and P = 0.04, respectively). Overall, pepperweed invasion had a net negative effect on ecosystem N status, depleting soil total N to potentially cause a negative feedback to invasion in the long term.