The molecular basis for selective assembly of the UBAP1-containing, endosome-specific ESCRT-I complex

ESCRT-I is essential for the multivesicular body (MVB) sorting of ubiquitinated cargo such as epidermal growth factor receptor, as well as for divergent cellular functions such as cell division and retroviral budding. ESCRT-I has four subunits; TSG101, VPS28, VPS37 and MVB12. There are several members of VPS37 and MVB12 families in mammalian cells, and their differential incorporation into ESCRT-I could provide function-specific variants of the complex. However, it remains unclear whether these different forms of VPS37 and MVB12 combine randomly or generate selective pairings within ESCRT-I, and what the mechanistic basis for such pairing would be. Here we show that the incorporation into ESCRT-I of two MVB12 members, UBAP1 and MVB12A, is highly selective with respect to their VPS37 partners. We map the selective assembly of UBAP1/VPS37A to the core ESCRT-I binding domain of VPS37A. In contrast, selective integration of UBAP1 requires both the minimal ESCRT-I binding region and a neighbouring predicted helix. The biochemical specificity in ESCRT-I assembly is matched by functional specialisation, since siRNA-mediated depletion of UBAP1, but not MVB12A or MVB12B, disrupts ubiquitin-dependent sorting at the MVB.