Stacking fault tetrahedra, the three-dimensional crystalline defects bounded by stacking faults and stair-rod dislocations, are often observed in quenched or irradiated face-centred cubic metals and alloys. All of the stacking fault tetrahedra experimentally observed to date are believed to originate from vacancies. Here we report, in contrast to the classical vacancyoriginated ones, a new kind of stacking fault tetrahedra formed via the interaction and crossslip of partial dislocations in gold nanocrystals. The complete atomic-scale processes of nucleation, migration and annihilation of the dislocation-originated stacking fault tetrahedra are revealed by in situ high-resolution observations and molecular dynamics simulations. The dislocation-originated stacking fault tetrahedra can undergo migration and annihilation due to mechanical loading in a manner that is not expected in bulk samples. These results uncover a unique deformation mechanism via dislocation interaction inside the confined volume of nanocrystals and have important implications regarding the size effect on the mechanical behaviour of small-volume materials.