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Related Concept Videos

Intralumenal Vesicles and Multivesicular Bodies01:38

Intralumenal Vesicles and Multivesicular Bodies

Intraluminal vesicles (ILVs) are small vesicles 50-80 nm in diameter formed during the maturation of early endosomes. A specialized endosome containing numerous ILVs is called a multivesicular body (MVB). ILVs contain internalized molecules such as antigens, nucleic acids, proteins, and metabolites. Some of these molecules are released from the MVBs inside exosomes and are transported to other cells. Other MVBs contain molecules that are retained in the ILVs and are later degraded within the...
Vesicular Tubular Clusters01:45

Vesicular Tubular Clusters

After budding out from the ER membrane, some COPII vesicles lose their coat and fuse with one another to form larger vesicles and interconnected tubules called vesicular tubular clusters or VTCs. These clusters constitute a compartment at the ER-Golgi interface known as ERGIC (Endoplasmic Reticulum Golgi Intermediate Compartment). The ERGIC is a mobile membrane-bound cargo transport system that sorts proteins secreted from ER and delivers them to the Golgi.
With the help of motor proteins such...
Overview of Protein Sorting and Transport01:45

Overview of Protein Sorting and Transport

Eukaryotic cells have different membrane-bound organelles with distinct protein requirements. The process by which proteins are targeted to a specific organelle is called protein sorting.
Protein sorting can be of two types: signal-based sorting and vesicle-based trafficking. In signal-based sorting, specific amino acid sequences called sorting signals target proteins to the proper location inside the cell either via gated transport or by protein translocation.  In gated transport, folded...
Transport Across the Golgi01:26

Transport Across the Golgi

While it is unclear how molecules move between adjacent Golgi cisternae, it is apparent that the molecules move from cis- cisterna, the entry face, to the trans- cisterna, the exit face. Experiments initially suggested vesicles that bud from one cisterna and fuse with the next cisterna to transport proteins between the cisternae. This vesicular transport model describes the Golgi apparatus as a relatively static structure with a unique enzyme composition in each cisterna. Molecules are...
Mitochondrial Protein Sorting01:39

Mitochondrial Protein Sorting

Mitochondria are double-membrane organelles of the eukaryotes involved in cellular metabolism, signaling, ATP synthesis, and programmed cell death.  Each of these processes requires specific proteins and enzymes that must be correctly sorted to the right mitochondrial subcompartment for the proper functioning of the organelle.
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COP Coated Vesicles00:59

COP Coated Vesicles

Membrane-enclosed structures called vesicles transport proteins and lipids across the cell. The vesicles derive their cargo from the plasma membrane, Golgi, ER, or endosome. Coated vesicles are spherical, protein-coated carriers with a 50–100 nm diameter that mediate bidirectional transport between the ER and the Golgi. The distribution of proteins between the ER and Golgi complex is dynamic and is maintained by different coated vesicles. Their formation is driven by the assembly of different...

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Related Experiment Video

Updated: Jun 19, 2026

Setting a Successful Sorting for Extracellular Vesicle Isolation
08:37

Setting a Successful Sorting for Extracellular Vesicle Isolation

Published on: October 11, 2024

Cargo sorting into multivesicular bodies in vitro.

John H Tran1, Ching-Jen Chen, Scott Emr

  • 1Department of Molecular and Cell Biology, and Howard Hughes Medical Institute, University of California, Berkeley, CA 94720, USA. tranj1@mskcc.org

Proceedings of the National Academy of Sciences of the United States of America
|October 7, 2009
PubMed
Summary
This summary is machine-generated.

Researchers developed a cell-free system to study how proteins enter endosomes. This method tracks the internalization of yeast carboxypeptidase S (CPS), revealing key protein requirements for this essential cellular process.

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Published on: April 12, 2024

Area of Science:

  • Cell Biology
  • Molecular Biology
  • Protein Trafficking

Background:

  • Multivesicular bodies (MVBs) are crucial for protein sorting and transport within eukaryotic cells.
  • Proteins involved in biosynthetic and endocytic pathways are internalized into MVBs.
  • Genetic studies have identified several proteins essential for MVB cargo internalization.

Purpose of the Study:

  • To develop a cell-free system to study the internalization of biosynthetic cargo into endosomes.
  • To investigate the molecular requirements for the internalization of yeast carboxypeptidase S (CPS).

Main Methods:

  • Developed a cell-free reaction using isolated yeast endosomes.
  • Utilized a recombinant CPS with a biotinylation tag.
  • Employed Escherichia coli biotin ligase for tagging cytosol-exposed CPS.
  • Quantified internalization by detecting trypsin-inaccessible, membrane-protected CPS.

Main Results:

  • The cell-free system successfully recapitulated CPS internalization into endosomes.
  • Internalization was observed in a vps27 yeast mutant blocked in MVB formation.
  • Biotinylated CPS internalization required ATP, functional Vps27p and Vps4p proteins.
  • The process depended on an exposed lysine residue for CPS ubiquitylation.

Conclusions:

  • The cell-free system provides a novel approach to study endosomal protein internalization.
  • Vps27p, Vps4p, ATP, and ubiquitylation are critical for CPS internalization into endosomes.
  • This system facilitates the dissection of protein trafficking machinery involved in MVB formation.