microRNA (miRNA) is a small non-coding RNA that plays a key role in the post-transcriptional regulation of genes. miRNAs influence various physiological processes and diseases such as cancer, heart disease and neurodegenerative disorders. Although miRNAs exist inside cells, they are also known to be packaged into extracellular vesicles (EV) such as exosomes and microvesicles, and thus to exist in the extracellular space as well.
The sorting (selection) of miRNAs into extracellular vesicles is an active process, and its mechanism has not yet been fully elucidated. However, several mechanisms have been proposed to explain how miRNAs are sorted into EVs.
Sorting dependent on sequence motifs:
Studies have shown that the presence of specific sequence motifs in a miRNA can drive its packaging into EVs. For example, the presence of ‘EXOmotifs’ or the ‘GGAG’ motif in a miRNA is associated with its selection into exosomes.
Sorting dependent on the miRNA-induced silencing complex (miRISC):
miRNAs associated with the miRISC, which induces gene silencing, may be selectively sorted into EVs. AGO2, a protein that is part of this complex, is found within EVs.
The miRNA-Induced Silencing Complex (miRISC) is the intracellular machinery through which miRNAs exert their gene-regulatory functions. This complex contains proteins such as Argonaute (AGO), which bind to miRNAs and guide them to target mRNAs.
Much remains to be elucidated about the mechanism by which miRISC-associated miRNAs are selectively sorted into EVs. However, it has been suggested that EVs, including exosomes, contain components of the miRISC such as AGO proteins. This means that miRNAs bound to these proteins may be co-packaged into EVs.
The precise molecular details of how such a phenomenon occurs are still under study. The presence of AGO2, a constituent protein of the miRISC, within EVs may play a role in this. There is also some evidence suggesting that the binding of miRISC and miRNA influences their distribution between the cell and the extracellular environment, but this requires further confirmation.
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 HnRNPs:
hnRNPs is an English abbreviation that stands for “heterogeneous nuclear ribonucleoproteins”. The full name in English is “Heterogeneous Nuclear Ribonucleoproteins”.
They support a series of RNA processing events, in particular the regulation of splicing, RNA transport, mRNA stability and translation.
hnRNPs are usually present in the cell nucleus, but they can also move into the cytoplasm. Their function is regulated mainly through interactions with RNA. hnRNPs have the ability to recognize and bind specific RNA motifs, which allows specific RNA molecules to be sent to specific intracellular sites.
With regard to the sorting of miRNAs, hnRNPs are thought to bind to miRNAs and play a role in guiding them into extracellular vesicles (EV). In particular, it has been suggested that the protein hnRNPA2B1 recognizes specific motifs of miRNAs and is sumoylated (a post-translational modification) in order to sort them into EVs.
However, the relationship between hnRNPs and miRNA sorting has not yet been fully elucidated, 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:
The sumoylated (a post-translational modification of proteins) hnRNPA2B1 protein may also be involved in the sorting of miRNAs. Sumoylated hnRNPA2B1 recognizes specific motifs of 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, and it affects various functions of the protein.
Specific changes include the following:
- Activation or inactivation of the protein: When SUMO binds to a protein, it can enhance or suppress the activity of that protein. This applies to a wide range of proteins, such as enzymes and transcription factors.
- Localization of the protein: Sumoylation can change the intracellular location of a protein. For example, a protein bound by SUMO may move more readily into the cell nucleus.
- Protein–protein interactions: When SUMO binds to a protein, it can change how that protein interacts with other proteins. This occurs either because the protein gains the ability to bind to new partners or because it loses its binding with existing partners.
- Stability of the protein: Sumoylation affects the stability of a protein and can extend or shorten its lifespan.
Through the effects described above, sumoylation plays an important role in regulating cellular processes such as cell division, DNA repair, transcriptional 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.
Sorting dependent on 4E-T:
4E-T (EIF4E transporter) is a type of RNA-binding protein that plays a role in controlling the post-transcriptional metabolism and translation of RNA.
EIF4E stands for “Eukaryotic Translation Initiation Factor 4E” and is a protein that plays a role in initiating the translation of proteins (the process of converting genetic information into proteins). 4E-T, on the other hand, transports this EIF4E and is involved in the regulation of translation.
Specifically, 4E-T binds to EIF4E and functions as a chaperone* for it. This allows EIF4E to move precisely to where it is needed and to fulfill its role at the appropriate time.
In addition, 4E-T also plays a role in controlling the selective translation of mRNAs. In particular, under cellular stress, 4E-T regulates the translation of mRNAs of specific stress-response proteins.
Furthermore, recent studies have shown that 4E-T has a role in sorting miRNAs into exosomes (a type of extracellular vesicle). This allows cells to control which miRNAs are included in 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 plays a role in helping proteins adopt their proper three-dimensional structure (that is, a correctly folded state). If a protein is not folded correctly, its function may be impaired or the cell may experience stress.
Chaperone proteins not only help newly synthesized proteins fold properly, but also help repair abnormally shaped proteins, and help degrade proteins that cannot be repaired. In this way, cells can maintain the quality of their proteins.
Chaperones are also involved in the transport of proteins. That is, they can play a role in carrying a protein to a specific location within the cell. In this case, the chaperone acts as a kind of “guide” for the protein and ensures that it reaches the correct location.
Reference: Hartl, F. U., Bracher, A., & Hayer-Hartl, M. (2011). Molecular chaperones in protein folding and proteostasis. Nature, 475(7356), 324-332.