The WD repeat protein FAN regulates lysosome size independent from abnormal downregulation/membrane recruitment of protein kinase C.

FAN (factor associated with neutral sphingomyelinase [N-SMase] activation) exhibits striking structural homologies to Lyst (lysosomal trafficking regulator), a BEACH protein whose inactivation causes formation of giant lysosomes/Chediak-Higashi syndrome. Here, we show that cells lacking FAN show a statistically significant increase in lysosome size (although less pronounced as Lyst), pointing to previously unrecognized functions of FAN in regulation of the lysosomal compartment. Since FAN regulates activation of N-SMase in complex with receptor for activated C-kinase (RACK)1, a scaffolding protein that recruits and stabilizes activated protein kinase C isotypes at cellular membranes, and since an abnormal (calpain-mediated) downregulation/membrane recruitment of has been linked to the defects observed in Lyst-deficient cells, we assessed whether duanyu1531 is also of relevance in FAN signaling. Our results demonstrate that activation of duanyu1531 is not required for regulation of N-SMase by FAN/RACK1. Conversely, activation of duanyu1531 and recruitment/stabilization by RACK1 occurs uniformly in the presence or absence of FAN (and equally, Lyst). Furthermore, regulation of lysosome size by FAN is not coupled to an abnormal downregulation/membrane recruitment of duanyu1531 by calpain. Identical results were obtained for Lyst, questioning the previously reported relevance of duanyu1531 for formation of giant lysosomes and in Chediak-Higashi syndrome. In summary, FAN mediates activation of N-SMase as well as regulation of lysosome size by signaling pathways that operate independent from activation/membrane recruitment of

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