ESCRT


The endosomal sorting complexes required for transport proteins are part of a pathway inside cells that helps sort and move other proteins. One of their main jobs is to form structures called multivesicular bodies which help sending of certain proteins, especially ones tagged for removal, to compartments in the cell called lysosomes where they get broken down.
The ESCRT system is made up of five separate cytosolic, peripheral membrane protein complexes, known as ESCRT-0, ESCRT-I, ESCRT-II, ESCRT-III and Vps4. Together with a number of accessory proteins, these ESCRT complexes enable a unique mode of membrane remodeling that results in membranes budding away from the cytoplasm. These ESCRT components have been isolated and studied in a number of organisms including yeast and humans.
The ESCRT machinery plays a vital role in a number of cellular processes including multivesicular body biogenesis and cytokinetic abscission. Multivesicular body biogenesis is a process in which ubiquitin-tagged proteins enter organelles called endosomes via the formation of vesicles. Cells break down damaged membrane proteins within two main complexes: the proteasome and the lysosome. A small tag called ubiquitin gets attached to them. The tag leads proteins to either the proteasome or the lysosome for destruction. For the lysosomal route the tagged proteins are sent into small compartments inside the cell called endosomes, specifically a kind called multivesicular bodies, MVBs are made when part of the endosome membrane folds inward and forms intralumenal vesicles. These intraluminal vesicles carry the proteins meant to be destroyed, and when an MVB joins with a lysosome, the vesicles and the proteins inside get broken down.
When autophagy does not work well like in cells with ESCRT mutations the cell cannot get rid of clumps of damaged proteins very well. These protein clumps are commonly seen in neurodegenerative disease like Alzheimer's or Parkinson's.
Cytokinetic abscission is the process where the intercellular bridge between two daughter cells is cut, completing cell division. In many animal cells, the ESCRT-III machinery is responsible for this process. The ICB is initially under high tension, which can prevent proper abscission in epithelial cells by interfering with the assembly of ESCRT-III.

ESCRT complexes and accessory proteins

The ESCRT system is like a team of cellular machines of ESCRT protein complexes and accessory proteins that help manage and remodel cell membranes.
The ESCRT complexes are needed for membrane-cutting events in the cell: MVBs are used to sort and recycle cell component, then viruses like HIV-1 bud off from the cell, and during the final steps of cell division when two cells physically separate.
Even though all these processes involve similar types of membrane shaping, they don't all need the same ESCRT complexes, For example the ESCRT-II complex is not needed for HIV-1 to bud from the cell and its exact role in cytokinesis is still unclear.
Each of the ESCRT complexes and accessory proteins has unique structures that enable distinct biochemical functions. A number of synonyms exist for each protein component of the ESCRT machinery, both for yeast and metazoans.
A summary table of all of these proteins is provided at the table beside.

ESCRT-0

The ESCRT-0 complex plays a vital role in the generation of multivesicular bodies by binding and clustering ubiquitinated proteins and/or receptors on the surface of a cell. The complex is then responsible for binding to a lipid on the endosomal membrane, which recruits these tagged proteins to the endosome. Once properly localized, these proteins are then taken into the endosome via vesicles, forming multivesicular bodies, and are eventually delivered to the lysosome where they are degraded. This process is essential as it is the major pathway for the degradation of damaged proteins that have passed through the Golgi. The components of the ESCRT-0 complex exist as follows:
The complex is a 1:1 heterodimer of Vps27 and Hse1. Vps27 and Hse1 dimerize through antiparallel coiled-coil GAT domains. Both Vps27 and Hse1 contain an amino-terminal VHS domain. These VHS domains bind the ubiquitin on proteins the cell aims to degrade. Ubiquitin can also associate with ubiquitin interacting motifs such as the one on Hse1 or the double-sided domain found on Vps27. A FYVE domain is found sandwiched between the VHS and ubiquitin interacting motif domains of Vps27. Phosphatidylinositol 3-phosphate, a common endosomal lipid, binds to this FYVE domain resulting in the recruitment of ESCRT-0 to the endosome. Research has made comprehensive analysis of the interaction between the ESCRT-0 complex and ubiquitin using isothermal titration calorimetry, this technique has been used for measures molecular binding strength. Research shows five main results:
  1. Ubiquitin Binding Behavior: Binding to the different ubiquitin-binding domains within ESCRT-0 is non-cooperative, meaning each domain interacts with ubiquitin independently.
  2. Binding Affinity: The Hrs subunit contains a double ubiquitin interacting motif. This DUIM has a binding affinity more than as twice as strong as the UBDs found in the STAM subunit and this indicates that Hrs in the primary ubiquitin binding protein within the ESCRT-0 complex.
  3. Localization in the Cell : Both Hrs and STAM are found on endosomal membranes.
  4. Complex Formation: By using atomic force microscopy, researchers observed that ESCRT-0 mainly forms: Heterodimers. Heterotetramers in the presence of lipid membranes. Hydrodynamic analysis of ESCRT-0 inside cells confirmed that it primarily exists as a heterotetramer
  5. Updated Functional Model: based on these results, the researchers proposed a revised model where ESCRT-0 plays a central role in recruiting and concentrating ubiquitinated cargo at the endosome for degradation.

    ESCRT-I

The role of the ESCRT-I complex is to assist in the generation of multivesicular bodies by clustering ubiquitinated proteins and acting as a bridge between the ESCRT-0 and ESCRT-II complexes. It also plays a role in membrane recognition and remodeling during membrane abscission by forming rings on either side of the midbody in dividing cells. ESCRT-I is also responsible for recruiting ESCRT-III, which forms the constriction zone just before the cells separate. Furthermore, ESCRT-I contributes to viral budding by interacting with specific viral proteins, leading to the recruitment of additional ESCRT machinery to the potential site of viral release.
The ESCRT-I complex is a heterotetramer composed of Vps23, Vps28, Vps37, and Mvb12. The assembled heterotetramer appears as a rod-shaped stalk formed by Vps23, Vps37, and Mvb12 with a fanned cap composed of single helices from Vps23, Vps28, and Vps37. Vps23 contains a ubiquitin E2 variant domain, which is responsible for binding of ubiquitin, the ESCRT-0 complex, and to the PTAP motif of viral Gag proteins. Immediately following this domain is a proline-rich motif, which directs ESCRT-I to the midbody during membrane abscission. Mvb12 can also bind ubiquitin via its carboxy-terminus. Vps28 facilitates the interaction between ESCRT-I and ESCRT-II by associating with the GLUE domain of Vps36 through its carboxy-terminal four-helix bundle domain.
Finally, the ESCRT-I complex co-assembles with ESCRT-II on membranes to form a 1:1 supercomplex that facilitates the budding of the limiting membrane of the multivesicular body into its lumen. Interestingly, in processes such as HIV-1 budding and cytokinesis where ESCRT machinery is not required for bud formation. ESCRT-I appears to function independently of ESCRT-II and is likely involved mainly in recruiting ESCRT-III. Structurally, ESCRT-I subunits heterotetramerize through two contiguous but distinct core regions. Vps23, Vps37, and Mvb12 form a 13 nm-long stalk that includes an unusual antiparallel coiled-coil structure. Meanwhile, Vps23, Vps28, and Vps37 contribute to a headpiece region composed of three pairs of antiparallel helices, arranged in a fan-like shape. Together, the stalk and headpiece form a single rigid structure approximately 18 nm in length.

ESCRT-II

The ESCRT-II complex plays a more critical role that ESCRT-I in multivesicular body biogenesis, as its overexpression can rescue the loss of ESCRT-I function in yeast, whereas the reverse is not true. Despite its central function in MVB formation and membrane budding, ESCRT is not essential for HIV-1 budding or cytokinesis. Unlike ESCRT-0 and ESCRT-I, ESCRT-II binds ubiquitinated cargo at a single site and acts as a key bridging complex between the upstream ESCRT machinery and the downstream ESCRT-III complex, which performs membrane scission. ESCRT-II may also be involved in linking MVBs to microtubules through interactions with RILP, Rab7, and dynein motor proteins.
Structurally, ESCRT-II forms a Y-shaped complex, with Vps22 and Vps36 at the base and two Vps25 subunits as the arms. Vps22 and Vps36 interact tightly through their winged-helix domains and are mutually required for folding and stability, while the Vps25 subunits are more loosely attached and do not interact with each other. The second WH domain of Vps25 mediates interaction with the ESCRT-III subunit Vps20, and both Vps25 mediates interaction with the ESCRT-III subunit Vps20, and both Vps25 copies are essential for proper function.
Additionally, the N-terminus of Vps22 contains a basic helix that aids in membrane targeting without lipid specificity, In yeast, Vps36 features a GLUE domain containing two inserted NpI4 Zinc fingers, which mediate binding to the Vps28 subunit of ESCRT-I and to ubiquitin, respectively. In mammals, the GLUE domain of VPS36 lacks these zinc fingers but can bind ubiquitin directly. Furthermore, the linker region between the GLUE and WH1 domains of yeast Vps36 may serve as a secondary binding site for ESCRT-I, a region that is conserved in the human homolog, although its role remains unconfirmed.
The ESCRT-II complex functions primarily during the biogenesis of multivesicular bodies and the delivery of ubiquitin tagged proteins to the endosome. Ubiquitin-tagged proteins are transferred sequentially from ESCRT-0 to ESCRT-I and then to ESCRT-II. ESCRT-II associates with ESCRT-III, which pinches off the cargo containing vesicle. The specific aspects of ESCRT-II are as follows:
ESCRT-II is a heterotetramer composed of two Vps25 subunits, one Vps22 subunit, and one Vps36 subunit. Vps25 molecules contain PPXY motifs, which bind to winged-helix motifs of Vps22 and Vps36 forming a Y-shaped complex with Vps22 and Vps36 at the base and the Vps25 subunits forming the arms. Vps25 molecules also contain WH motifs that mediate the interaction between ESCRT-II and ESCRT-III. Vps36 includes a GLUE domain that binds phosphatidylinositol 3-phosphate and Vps28 subunit of ESCRT-I. Two zinc finger domains are looped into the GLUE domain of yeast Vps36. One of these zinc finger domains binds the carboxy-terminal domain of Vps28 while the other interacts with ubiquitin.