Hierarchical 3D flower-like MoS2 nanospheres with the diameter of about 300 nm were successfully synthesized via a facile hydrothermal process assisted with CTAB. And we found that CTAB played a vital role on the formation of MoS2 architectures. As the concentration of CTAB increasing, MoS2 nanopowders with sheet-like, flower-like, large size spherical morphologies were synthesized. And particle sizes of synthesized powders also increased. The optimum concentration of CTAB for the well-defined 3D hierarchical spherical morphology of MoS2 was 6 g/L. In addition, a feasible formation mechanism of the morphology evolution was put forward in detail on the basis of the experimental results. The gas-sensing properties of flower-like MoS2 nanospheres were tested. The result indicated that the sensor exhibited high response and selectivity towards NO2, which may be related to the open and well-defined structures and the abundant micro reaction rooms assembled by ultrathin nanosheets.