Interaction of furin in immature secretory granules from neuroendocrine cells with the AP-1 adaptor complex is modulated by casein kinase II phosphorylation

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Abstract
The composition of secretory granules in neuroendocrine and endocrine cells is determined by two sorting events; the first in the trans‐Golgi complex (TGN), the second in the immature secretory granule (ISG). Sorting from the ISG, which may be mediated by the AP‐1 type adaptor complex and clathrin‐coated vesicles, occurs during ISG maturation. Here we show that furin, a ubiquitously expressed, TGN/endosomal membrane endoprotease, is present in the regulated pathway of neuroendocrine cells where it is found in ISGs. By contrast, TGN38, a membrane protein that is also routed through the TGN/endosomal system does not enter ISGs. Furin, however, is excluded from mature secretory granules, suggesting that the endoprotease is retrieved from the clathrin‐coated ISGs. Consistent with this, we show that the furin cytoplasmic domain interacts with AP‐1, a component of the TGN/ISG‐localized clathrin sorting machinery. Interaction between AP‐1 and furin is dependent on phosphorylation of the enzyme's cytoplasmic domain by casein kinase II. Finally, in support of a requirement for the phosphorylation‐dependent association of furin with AP‐1, expression of furin mutants that mimic either the phosphorylated or unphosphorylated forms of the endoprotease in AtT‐20 cells demonstrates that the integrity of the CKII sites is necessary for removal of furin from the regulated pathway.