Nitrogen (N) deficiency is one of the major constraints for potato (Solanum tuberosum L.) production, due to its shallow root system and poor capabilities of N mobilization from the deeper layers of the soil. The aim of this study was to elucidate plasma membrane (PM) proteins potentially involved in the tolerance towards N deficiency. We compared the PM proteome of the two potato cultivars ‘Lambada’ and ‘Topas’, contrasting in their N deficiency response under in vitro conditions. Using mass spectrometry we identified 65 proteins, which were differentially abundant in the two cultivars when submitted to N deficiency. Amongst those, candidate proteins were selected based on their potential to be involved in N transport under deficient conditions. Two members of the aquaporin family, StPIP1;1 and StPIP1;3, were studied in more detail. By means of a yeast growth assay we showed that these proteins do not facilitate transport of ammonia. The Arabidopsis (Arabidopsis thaliana) knockout mutants AtPIP1;1 and AtPIP1;3 showed a reduced N content and accumulation of anthocyanins under N deficiency. We conclude that PIP1;1 and PIP1;3 are indirectly involved in the N uptake under N deficient conditions and contribute to an increased N deficiency tolerance.