SummaryAlthough exocytosis in fungal cells takes place at hyphal tips, there also seems a line of circumstantial evidence suggesting the occurrence of exocytosis at other sites of cells, such as septa. To investigate whether exocytosis takes place at fungal septa, we monitored dynamics of EGFP-fused a-amylase (AmyB-EGFP), the representative secretory enzyme of the filamentous fungus Aspergillus oryzae. We found that AmyB-EGFP accumulates in Spitzenkörper at hyphal tips as well as septal periplasm between the plasma membrane and cell walls. The septal accumulation of AmyB-EGFP was a rapid process, and required microtubules but not F-actin. Thus, this process is independent of exocytosis at hyphal tips that requires both microtubules and F-actin. In addition, fluorescence recovery after photobleaching (FRAP) analysis of EGFP-fused AoSnc1 revealed that secretory vesicles constitutively fuse with the septal plasma membrane. These results demonstrated that exocytosis takes place at septa in addition to hyphal tips. Analysis of two plasma membrane transporters, AoUapC and AoGap1, revealed that they preferentially accumulate at septa and the lateral plasma membrane with no clear accumulation at apical Spitzenkörper, suggesting that non-tip directed exocytosis is important for delivery of these proteins.