“The siRNA or small interfering RNA is a 22 to 25 basepair long smaller molecules of RNA having a dinucleotide overhang at the 3’, interfere in the protein synthesis by blocking the translation.”


 The presence of the double-stranded RNA in a cell is a sign of danger for it. As the main three types of RNA in us are single-stranded.

The dsRNA is not found in our cells because our genetic material is DNA not RNA. the dsRNA is genetic material of some retroviruses thus, 

If dsRNA is present, it is a sign of infection, this will infect the cells too and can cause cell death. 

Note: the ribosomal DNA contains some amount of dsRNA and some hairpin RNA molecules too. 

The RNA is a type of nucleic acid which is present in the nucleus of a cell. Although it is not a genetic material in eukaryotes, some of the viruses known as retroviruses have RNA as their genetic material. 

The DNA is genetic material in all eukaryotes and prokaryotes, except retroviruses. 

The RNA is a ribonucleic acid made up of the ribose sugar instead of deoxyribose sugar of the DNA. 

For more detail, on RNA you can read our article on RNA: RNA: Structure and Function

First, let me brief you the RNA;

The RNA is ribonucleic acid, tRNA, rRNA and mRNA are three different types of RNA present in a cell. 

The tRNA is a transfer RNA helps in transferring the information for protein synthesis, the rRNA is a ribosomal RNA located in the ribosome, read the order of amino acids and the mRNA is a messenger RNA. 

The mRNA or messenger RNA has all the information for coding a particular protein

Functionally, mRNA is transcribed from the DNA and translated into the protein via the pathway of translation. 

The shRNA, miRNA and siRNA also some of the subsidiary type of RNA present in a minor amount for the regulation of gene expression. 

In the present article, we will learn the siRNA, its signification and its clinical applications. Trust me this article will clear your fundamentals on siRNA. 

So let’s start,

What is siRNA? 

The smaller double-stranded piece of RNA having a dinucleotide overhang at the 3’ end which functionally, degrade the mRNA and prevent the protein synthesis are siRNAs.

The siRNA also known as small interfering ribonucleic acid or silencing RNA is a molecule prevents the gene expression. 

The entire process of gene silencing through the siRNA is called a mechanism of RNA interference or siRNA knockdown.  

 The siRNA is functionally and structurally different from the other type of RNAs. 

Generally, other RNAs are single-stranded and made up of the long polynucleotide chain. 

On the other side, 

The siRNA is double-stranded, short and 20 to 25 nucleotides long. 

The source of siRNA is exogeneous.

Functionally, it blocks gene expression. 

Apart from all these, one of the unique characters of the siRNA is the presence of the 3’ OH dinucleotide overhang. See the figure,

The structure of siRNA having a guided strand, passanger strand and the dinucleotide overhang at the 3 end.

The structure of siRNA having a guided strand, passenger strand and the dinucleotide overhang at the 3 ends.

The siRNA is a double-stranded structure in which one strand is known as the guide strand and another strand is called the passenger strand. It is also called a sense strand and antisense strand, respectively.

In the year 1999, David Baulcombe and coworkers explained the role of the siRNA in the post-transcriptional modification. 

The function of siRNA: 

The main function of siRNA is to protect the cell from the exogenous mRNA attacks.

Functionally, the siRNA degrades the growing mRNA (exogenous as well as endogenous) and stop gene expression. 

The origin of the siRNA is exogeneous, it came from the viral infections. 

The eukaryotic cells have a very good responsive defence system called RNA interference. 

Now let us understand the entire mechanism in detail, 

RNA interference often denoted as RNAi is a biological process for mRNA degradation and subsequent gene silencing. 

In 1998, Fire and Mello unfold the mechanism of RNA interference. The role of the siRNA in the RNA interference was discovered in 1999. 

Once the retrovirus infects a cell it inserts its dsRNA into our cell. 

The specialized protein called dicer having a tetrameric manganese ions cuts or cleaves the dsRNA into smaller pieces. 

A special type of RNase, the dicer cleaves the RNA is a fashion that produces the dinucleotide overhang. 

This smaller fragments of dsRNA then incorporated into the protein complex having multiple subunits and form the RNAi-induced silencing complex, RISC. 

The RISC finds the appropriate mRNA target and cleaved it by a combination of endo and exonuclease activity. 

These smaller dsRNAs are ~22 to 25 basepair long, called the small interfering RNA or siRNA. 

The siRNA also have the phosphate group at the 5’ end of it. 

Also as we discussed, it has the dinucleotide overhang. It is believed that the overhang of dinucleotides is originated due to the activity of manganese ions present in the dicer. 

The guided strand of siRNA guide the protein complex to find the complementary dsRNA sequence present in a cell, once it recognized, it is cleaved and destroyed.

In this way, the natural RNA interference defence mechanism defends the cell from the viral infection via the siRNA.

Similarly, it can also able to destroy the mRNA of us by finding the complementary mRNA thus it modifies the properties of the chromosome by altering the epigenetic profile of the genome. 

 Applications of siRNA: 

The present mechanism is actively present in almost all eukaryotes and works against viral infections. 

Nowadays, scientists are using this knowledge for gene silencing and stopping gene expression for therapeutic uses.

Scientists are now synthesizing the artificial siRNA molecules specific to the mRNA of a gene they wish to block. 

By using the viral vector-based on nonviral-vector based artificial methods of transfer, the siRNA can be inserted into the cell. 

Read more on viral vector-based and nonviral- vector-based gene delivery: Gene Therapy: Types, Vectors [Viral and Non-Viral], Process, Applications and Limitations

The targetted mRNA is destroyed and the protein synthesis is regulated by this mechanism. 

Scientists are now trying to use the siRNA mediated gene silencing method for cancer-causing genes. 

The siRNA mediated method is used in the gene knockout and gene knockdown method for suppressing the gene expression.

It is used in the target validation. 

It is also used in pathway analysis and pathway identifications like cytokinesis, insulin signalling and cell defence mechanism etc.

Further, It is applicable in the gene redundancy study and gene functional studies. 

The carbon-based and non-carbon-based nanoparticle-mediated siRNA therapy is used in the drug delivery to the brain.

Interestingly, in 2001, Elbashir et al used the synthetic siRNA molecule for artificial gene silencing. 

Challenges in the siRNA: 

The RNA interference by siRNA is a novel approach, researchers are not much aware of how to use it, numerous problem associated with the use of siRNA in therapeutics,

The nuclease present in the plasma and the tissue degrades the foreign siRNA oligo molecules, however, the siRNA via nanoparticles shown some promising results, as we stated in the above section. 

Further, the effect of the present therapy is fewer and tissue-specific thus it is restricted to the localized sites. 

Because of its larger size, it is very difficult to pass it through the cell membrane, although nano-vectors can transfer the siRNA effectively. 

Off targeting of the siRNA is one of the major challenges is the siRNA research as it also degrades other mRNAs too. 

It is very difficult to use the siRNA for the therapeutic applications now, because of these challenges, notwithstanding, it will be applicable in future. 

Example of therapeutic siRNA:

SM2181 AUCUGAAGAAGGAGAAAAATT UCCUUUCUUUCUUUCGAAUTT 2% mRNA inhibition 0.3 nM

 

SM2172 AUCUGAAGAAGGAGAAAAATT UUUUUCUCCUUCUUCAGAUTT 88 % target mRNA inhibition 0.3 nM

Now, these are the two examples of the siRNA with the sense strand and the antisense strand, one with the higher activity and one with the lowest activity. Also, the table shows the concentration of siRNA. 

The data for the siRNA is now available on siRNAmod.

Related articles:

  1. DNA story: The structure and function of DNA
  2. RNA: Structure and Function

Conclusion: 

siRNA mediated therapies are one of the most promising tools for the biopharmaceutical fields. The specificity is one of the major obstacles in recent days, albeit, it can be a diagnostic tool for life treating disorders like cancer in future. 

Though viral and non-viral vectors such as lipid-based, peptide-based, oligo-based and polymer-mediated delivery systems are now available for siRNA, every method has some limitations. 

Resources:

  1. Dana H, Chalbatani GM, Mahmoodzadeh H, et al. Molecular Mechanisms and Biological Functions of siRNA. Int J Biomed Sci. 2017;13(2):48–57.
  2. The molecule of the month; Small interfering RNA.