The system for transporting RNA into Extracellular Vesicles (Exosomes)

Extracellular vesicles

MicroRNAs (miRNAs) are small non-coding RNAs that play a crucial role in the post-transcriptional regulation of genes. They influence various physiological processes and diseases, including cancer, heart disease, and neurodegenerative disorders. miRNAs exist inside cells but can also be found in the extracellular space packaged within extracellular vesicles (EVs), such as exosomes and microvesicles.

The sorting of miRNAs into extracellular vesicles is an active process, and its mechanism is not yet fully understood. However, several mechanisms have been proposed to explain how miRNAs are sorted into EVs.

Sorting dependent on sequence motifs:

Research has shown that the presence of specific sequence motifs in miRNAs can induce packaging into EVs. For instance, the presence of ‘EXOmotifs’ or ‘GGAG’ motifs in miRNAs is associated with sorting into exosomes.

Sorting dependent on the miRNA-induced silencing complex (miRISC):

miRNAs associated with the gene-silencing miRISC may be selectively sorted into EVs. The AGO2 protein, a part of this complex, is found within EVs.

The miRNA-Induced Silencing Complex (miRISC) is a cellular mechanism through which miRNAs exert their gene-regulating functions. This complex includes proteins such as Argonaute (AGO), which bind to miRNAs and guide them to target mRNAs.

While the mechanisms of miRISC-associated miRNA sorting into EVs remain largely unexplored, it has been suggested that components of miRISC, including AGO proteins, exist within EVs, including exosomes. It is possible that miRNAs bound to these proteins could be co-packaged into EVs.

The precise molecular details of how such phenomena occur are still under investigation. The presence of the miRISC constituent protein AGO2 within EVs may be playing a role. Additionally, some evidence suggests the binding of miRISC and miRNAs might affect their distribution between the cell and the extracellular environment, but further validation is needed.

Reference: Gibbings, D. J., Ciaudo, C., Erhardt, M., & Voinnet, O. (2009). Multivesicular bodies associate with components of miRNA effector complexes and modulate miRNA activity. Nature cell biology, 11(9), 1143-1149.

Please note that as the field of miRNA and EV biology is rapidly evolving, it’s possible that more recent research may provide additional insights into these mechanisms beyond my knowledge cutoff in September 2021.

Sorting dependent on Heterogeneous Nuclear Ribonucleoproteins (hnRNPs):

hnRNPs are abbreviations for the English term “Heterogeneous Nuclear Ribonucleoproteins”.

They support a series of RNA processing events, especially splicing, RNA transport, mRNA stability, and translation regulation.

While hnRNPs typically reside in the cell nucleus, they can also move to the cytoplasm. Their function is primarily regulated by interactions with RNA. hnRNPs have the ability to recognize and bind to specific RNA motifs, which allows specific RNA molecules to be transported to specific locations within the cell.

In terms of miRNA sorting, hnRNPs are thought to play a role by binding to miRNAs and guiding them into extracellular vesicles (EVs). In particular, it has been suggested that a protein called hnRNPA2B1 recognizes specific motifs in miRNAs and is Sumoylated (a type of post-translational modification) to sort them into EVs.

However, the relationship between hnRNPs and miRNA sorting is not fully understood, and further research is needed.

Reference: Santangelo, L., Giurato, G., Cicchini, C., Montaldo, C., Mancone, C., Tarallo, R., … & Weisz, A. (2016). The RNA-Binding Protein SYNCRIP Is a Component of the Hepatocyte Exosomal Machinery Controlling MicroRNA Sorting. Cell reports, 17(3), 799-808.

Sorting dependent on Sumoylated-HnRNPA2B1:

Sumoylated hnRNPA2B1, a post-translational modification of the protein, could also be involved in miRNA sorting. The Sumoylated hnRNPA2B1 recognizes specific motifs in miRNAs and sorts them into EVs.

Sumoylation is a chemical change that occurs when a protein called SUMO (Small Ubiquitin-like Modifier) binds to a protein, affecting various functions of the protein.

The specific changes include the following:

  1. Protein activation or deactivation: When SUMO binds to a protein, it can enhance or inhibit the activity of that protein. This applies to various proteins, such as enzymes, transcription factors, and others.
  2. Protein localization: Sumoylation may change the intracellular position of proteins. For example, proteins that SUMO binds to may be more likely to move to the cell nucleus.
  3. Protein-protein interaction: When SUMO binds to a protein, it can change how that protein interacts with other proteins. This occurs through the protein gaining the ability to bind with new partners or losing its connection with existing partners.
  4. Protein stability: Sumoylation can affect the stability of proteins and may extend or shorten their lifespan.

Through such impacts, Sumoylation plays a crucial role in regulating cellular processes like cell division, DNA repair, transcription control, and apoptosis (programmed cell death).

Reference: Villarroya-Beltri, C., Gutiérrez-Vázquez, C., Sánchez-Cabo, F., Pérez-Hernández, D., Vázquez, J., Martin-Cofreces, N., … & Falcón-Pérez, J. M. (2013). Sumoylated hnRNPA2B1 controls the sorting of miRNAs into exosomes through binding to specific motifs. Nature communications, 4(1), 1-10.

4E-T-dependent Sorting:

4E-T (EIF4E transporter) is a type of RNA-binding protein that plays a role in controlling the metabolism and translation of RNA after transcription.

EIF4E, short for “Eukaryotic Translation Initiation Factor 4E,” is a protein that initiates the translation of proteins (the process of converting genetic information into proteins). On the other hand, 4E-T transports this EIF4E and is involved in regulating translation.

Specifically, 4E-T binds with EIF4E and functions as a chaperone*. This allows EIF4E to accurately move to the necessary locations and perform its role at the appropriate timing.

Additionally, 4E-T also plays a role in controlling the selective translation of mRNA. Particularly under cellular stress, 4E-T regulates the translation of mRNA of specific stress response proteins.

Moreover, recent studies have shown that 4E-T has a role in sorting miRNA into exosomes (a type of extracellular vesicle). This allows cells to control which miRNA is included in the exosomes.

Reference: Kouhkan, F., Hafizi, M., Mobarra, N., Mossahebi-Mohammadi, M., Mohammadi, S., Behmanesh, M., … & Sattari, M. (2015). miRNAs: a new method for erythroid differentiation of hematopoietic stem cells without the presence of growth factors. Applied biochemistry and biotechnology, 175(2), 1134-1148.


What is a chaperone*?
A chaperone is a type of protein that helps other proteins attain their proper three-dimensional structures (that is, correctly folded states). If a protein is not correctly folded, its function may be impaired, or the cell may be stressed.

Chaperone proteins not only help newly synthesized proteins fold correctly, but they also help repair abnormally shaped proteins and help degrade unrepairable proteins. This allows cells to maintain protein quality.

Additionally, chaperones are also involved in protein transportation. That is, they can play a role in transporting proteins to specific locations within a cell. In this case, chaperones play a “guiding” role, ensuring that the protein reaches the correct destination.

Reference: Hartl, F. U., Bracher, A., & Hayer-Hartl, M. (2011). Molecular chaperones in protein folding and proteostasis. Nature, 475(7356), 324-332.