Interactions of ubiquitin and CHMP5 with the V domain of HD-PTP reveals role for regulation of Vps4 ATPase

Abstract

The family of Bro1 proteins coordinates the activity of the Endosomal Sorting Complexes Required for Transport (ESCRTs) to mediate a number of membrane remodeling events. These events culminate in membrane scission catalyzed by ESCRT-III, whose polymerization and disassembly is controlled by the AAA-ATPase, Vps4. Bro1-family members Alix and HD-PTP as well as yeast Bro1 have a central ‘V’ domains that non-covalently bind Ub and connect ubiquitinated proteins to ESCRT-driven functions such as the incorporation of ubiquitinated membrane proteins into intralumenal vesicles of multivesicular bodies. Recently, it was discovered that the V domain of yeast Bro1 binds the MIT domain of Vps4 to stimulate its ATPase activity. Here we determine the structural basis for how the V domain of human HD-PTP binds ubiquitin. The HD-PTP V domain also binds the MIT domain of Vps4 and ubiquitin-binding to the HD-PTP V domain enhances its ability to stimulate Vps4 ATPase activity. Additionally, we found V domains of both HD-PTP and Bro1 bind CHMP5 and Vps60, respectively, providing another potential molecular mechanism to alter Vps4 activity. These data support a model whereby contacts between ubiquitin, ESCRT-III, and Vps4 by V domains of the Bro1 family may coordinate late events in ESCRT-driven membrane remodeling events.

Nonlytic cellular release of hepatitis A virus requires dual capsid recruitment of the ESCRT-associated Bro1 domain proteins HD-PTP and ALIX

“…Although a role in eHAV release remains to be studied, NEDD4-family E3 ligases like ITCH are involved in the budding of many conventional enveloped viruses, including HIV-1 [47, 61, 62]. Ubiquitylation is a common signal for cargo to be trafficked into MVE, and the V domains of both ALIX and HD-PTP have specific ubiquitin-binding activities [42, 63, 64].…”

Bro1 family proteins harmonize cargo sorting with vesicle formation

“…V domain Proline Rich Region (PRR) ALIX LBPA, [77] CHMP4 [78] Ub, [22] GAG, [19] Syntenin, [18] AMSH3 [15] TSG101, [19,78] CEP55, [79] ALG2 [80] HD-PTP STAM2, CHMP4 [81] UBAP1, [82] VPS4, Spastin, Rps6KC1, SNX15, Ub, CHMP5, [5] UBPY [16] TSG101 [81] Bro1 Snf7 [83] Vps4, [13] Rfu1, [84] Ub, Vps60 [4] Doa4, [14] Vps23, [85] Rsp5 [85] Vps23/TSG101 as well as de-ubiquitinating enzymes such as Doa4, AMSH3, and UBPY, and the Ub-ligase Rsp5 (Table 1). [1,[14][15][16][17][18][19] Recruitment of Doa4 to the site of MVB cargo via Bro1 recycles Ub to escape degradation in the lysosome/vacuole, [14] and its association with the HECT Ub ligase Rsp5 may help reinforce cargo ubiquitination in a manner akin to that proposed for other Ub-cargo receptors.…”

“…[20] Bro1 is also an endosomal Ub-sorting receptor, that operates in parallel with other ESCRT-mediated Ub-cargo sorting pathways. [4,6] ALIX and HD-PTP harness ESCRT function during viral budding and MVB sorting in mammalian cells and bind Ub via their V domains, [5,7,19,21,22] yet the full physiological role that Ub-binding fulfills is likely incompletely understood. Recent work has shown that V domains of Bro1 and HD-PTP stimulate Vps4 ATPase activity and do so in a manner that is enhanced when those V domains bind Ub.…”

“…Bro1 family members have been proposed to act as cargo receptors based on the ability of their 'V' domains to bind Ub and other peptide motifs that drive sorting of cargoes into MVBs. [4][5][6][7][8][9] The N-terminal Bro1 Domain (BOD) of Bro1-family members binds ESCRT-III subunits (e.g., Snf7/CHMP4), an interaction thought to promote polymerization of ESCRT-III. [10][11][12] Recent work has also shown Bro1 family proteins can also regulate Vps4 activity directly though the binding of their V domain to Vps4, supporting a model where Bro1 family members coordinate ILV formation with the upstream events such as cargo incorporation.…”

“…[10][11][12] Recent work has also shown Bro1 family proteins can also regulate Vps4 activity directly though the binding of their V domain to Vps4, supporting a model where Bro1 family members coordinate ILV formation with the upstream events such as cargo incorporation. [5,13] TA B L E 1 Interactions of Bro1 family proteins during MVB sorting, organized by domains…”