GM-CSF is critical for dendritic cell (DC) survival and differentiation in vitro. To study its effect on DC development and function in vivo, we used a gene transfer vector to transiently overexpress GM-CSF in mice. We found that up to 24% of splenocytes became CD11c+ and the number of DC increased up to 260-fold to 3 × 108 cells. DC numbers remained substantially elevated even 75 days after treatment. The DC population was either CD8α+CD4− or CD8α−CD4− but not CD8α+CD4+ or CD8α−CD4+. This differs substantially from subsets recruited in normal or Flt3 ligand-treated mice or using GM-CSF protein injections. GM-CSF-recruited DC secreted extremely high levels of TNF-α compared with minimal amounts in DC from normal or Flt3 ligand-treated mice. Recruited DC also produced elevated levels of IL-6 but almost no IFN-γ. GM-CSF DC had robust immune function compared with controls. They had an increased rate of Ag capture and caused greater allogeneic and Ag-specific T cell stimulation. Furthermore, GM-CSF-recruited DC increased NK cell lytic activity after coculture. The enhanced T cell and NK cell immunostimulation by GM-CSF DC was in part dependent on their secretion of TNF-α. Our findings show that GM-CSF can have an important role in DC development and recruitment in vivo and has potential application to immunotherapy in recruiting massive numbers of DC with enhanced ability to activate effector cells.