Formation of secretory organelles requires the coupling of cargo selection to targeting into the correct exocytic pathway. While the assembly of regulated secretory granules is driven in part by selective aggregation and retention of content, we recently reported that AP-1 recruitment of clathrin is essential to the initial formation of WPB at the TGN. A selective coaggregation process might include recruitment of components required for targeting to the regulated secretory pathway. However, we find that acquisition of the regulated secretory phenotype by Weibel-Palade bodies (WPB) in endothelial cells is coupled to but can be separated from formation of the distinctive granule core by ablation of the AP-1 effectors aftiphilin and gamma-synergin. Their depletion by siRNA leads to WPB that fail to respond to secretagogue and release their content in an unregulated manner. We find that these nonresponsive WPB have density, markers of maturation, and highly multimerized von Willebrand factor (VWF) similar to those of wild-type granules. Thus, by also recruiting aftiphilin/gamma-synergin in addition to clathrin, AP-1 coordinates formation of WPB with their acquisition of a regulated secretory phenotype.