A highly conserved nuclear exosome complex, which contains 3’-5’ exonuclease activity, plays a critical role in processing various types of RNAs and degrading unstable and aberrantly transcribed RNAs. Here, we discover the role of the nuclear exosome in regulating the positioning of cohesin at highly transcribed convergent genes. We use in situ Hi-C to show that the nuclear exosome regulates intra-chromosomal interactions and the formation of interacting domain boundaries through cohesin regulation. However, the exosome-coupled transcription termination does not affect the chromatin 3D organization. Instead, we found a close correlation between the cohesin positioning and the defective transcription elongation observed in the exosome-depleted cells. Altogether, these results suggest the novel functions of nuclear exosome in proper positioning of cohesin, the maintenance of chromatin 3D organization, and gene expression.