Centrosomal P4.1-associated protein is a novel regulator of ESCRT pathway function during endosome maturation

Previously, we reported that centrosomal P4.1-associated protein (CPAP/CENPJ/SAS-4) positively regulates multivesicular body (MVB) biogenesis and endocytic vesicular transport (EVT). Here, we show that CPAP is required for Rab5-to-Rab7 conversion and that the recruitment of TSG101 to early endosomes (EEs) is the molecular mechanism by which CPAP promotes MVB formation. CPAP depletion disrupts Rab7 and TSG101 recruitment to EEs and blocks endosome maturation. While endogenous CPAP co-precipitates with the ESCRT-proteins TSG101, HRS, and ALIX, exogenously expressed CPAP interacts only with TSG101. CPAP localizes to the endosomes and co-localizes with HRS and TSG101 during EVT progression. TSG101 recruitment to the endosomes, Rab5-to-Rab7 conversion, and EVT of EGFR to MVB in CPAP-depleted cells are restored by re-introduction of CPAP or overexpression of, but not endogenous, HRS. These observations show that CPAP is an ESCRT-associated protein, which functions upstream of TSG101, but in parallel to HRS, during MVB formation and EVT of cargo to the lysosomes.