Although the presence of nanoplastics in aquatic and terrestrial ecosystems has received increasing attention, little is known about its potential effect on ecosystem processes and functions. Here, we evaluated if differentially charged polystyrene (PS) nanoplastics (PS-NH2 and PS-SO3H) exhibit distinct influences on microbial community structure, nitrogen removal processes (denitrification and anammox), emissions of greenhouse gases (CO2, CH4, and N2O), and ecosystem multifunctionality in soils with and without earthworms through a 42-day microcosm experiment. Our results indicated that nanoplastics significantly altered soil microbial community structure and potential functions, with more pronounced effects for positively charged PS-NH2 than for negatively charged PS-SO3H. Ecologically relevant concentration (3 g kg−1) of nanoplastics inhibited both soil denitrification and anammox rates, while environmentally realistic concentration (0.3 g kg−1) of nanoplastics decreased the denitrification rate and enhanced the anammox rate. The soil N2O flux was always inhibited 6%–51% by both types of nanoplastics, whereas emissions of CO2 and CH4 were enhanced by nanoplastics in most cases. Significantly, although N2O emissions were decreased by nanoplastics, the global warming potential of total greenhouse gases was increased 21%–75% by nanoplastics in soils without earthworms. Moreover, ecosystem multifunctionality was increased 4%–12% by 0.3 g kg−1 of nanoplastics but decreased 4%–11% by 3 g kg−1 of nanoplastics. Our findings provide the only evidence to date that the rapid increase in nanoplastics is altering not only ecosystem structure and processes but also ecosystem multifunctionality, and it may increase the emission of CO2 and CH4 and their global warming potential to some extent.