Question 1

What are intraluminal vesicles and how do they form inside endosomes?

Accepted Answer

Intraluminal vesicles (ILVs) are small vesicles 50-80 nm in diameter formed when the endosomal membrane bends inward during early endosome maturation. ESCRT protein complexes coordinate this inward budding process, with phosphatidylinositol 3-phosphate acting as a docking site for ESCRTs. Ubiquitin-tagged proteins are recognized and sorted into these ILVs by sequential ESCRT complexes.

Question 2

What is a multivesicular body and what does it contain?

Accepted Answer

A multivesicular body (MVB) is a specialized endosome containing numerous intraluminal vesicles, measuring 400-500 nm in diameter. MVBs are spherical organelles that represent an intermediate form between early and late endosomes. They contain internalized molecules such as antigens, nucleic acids, proteins, and metabolites that are either released via exosomes or retained for degradation.

Question 3

How do ESCRT complexes work to form intraluminal vesicles?

Accepted Answer

Four types of ESCRT complexes act sequentially during ILV formation. ESCRT-0 recognizes ubiquitinated proteins on the endosomal membrane, while ESCRT-I and II induce inward membrane budding. ESCRT-II triggers homo-oligomerization of ESCRT-III, which recruits deubiquitinating enzymes and works with the ATPase Vps4 to mediate membrane fission and ILV release.

Question 4

Why are proteins ubiquitinated before being sorted into intraluminal vesicles?

Accepted Answer

Ubiquitin tagging marks proteins destined for degradation or signaling proteins that must be sequestered from the cytosol. E3 ligases attach ubiquitin to these proteins at the plasma membrane during receptor-mediated endocytosis. ESCRT complexes specifically recognize ubiquitin-tagged proteins, ensuring selective sorting into ILVs for proper cellular trafficking and degradation.

Question 5

How do retroviruses hijack the MVB formation machinery?

Accepted Answer

Enveloped retroviruses like HIV exploit ESCRT complexes and Vps4 for viral particle release. HIV codes for ubiquitinated proteins such as Gag, NC-p2, and Tat that are recognized and sorted into viral buds by ESCRTs. Once viral particles bud from infected cells, they are released into extracellular space to infect other host cells, mimicking normal ILV formation.

Question 6

What is the role of phosphatidylinositol 3-phosphate in intraluminal vesicle formation?

Accepted Answer

Phosphatidylinositol 3-phosphate (PtdIns3P) is required for ILV formation and acts as a docking site for ESCRT complexes on the endosomal membrane. This lipid modification enables ESCRTs to recognize and bind to the membrane, facilitating the inward bending necessary for ILV budding and subsequent membrane fission.

Question 7

What happens to molecules inside multivesicular bodies after formation?

Accepted Answer

Molecules within MVBs follow two distinct pathways. Some are released from MVBs inside exosomes and transported to other cells for intercellular communication. Other MVBs retain their molecular cargo within ILVs, which are later degraded within the cell through lysosomal fusion, enabling cellular waste removal and nutrient recycling.

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