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In this article we will discuss about:- 1. Introduction to Micro RNAs 2. Formation of Micro RNAs 3. Processing 4. Differences Between Small Interfering RNA (siRNA) and Micro RNA (miRNA)5. Roles.
Introduction to Micro RNAs (miRNA):
Micro RNAs were first described for the worm C. elegans in 1993 by Lee and colleagues in the Victor Ambros lab. However, the term micro RNA was only introduced in 2001. By 2002, miRNAs have been confirmed in various plants and animals, including C. elegans, human and the plant Arabidopsis thaliana.
Work at the University of Louisville has resulted in the production of microarrays containing all known miRNAs for human, mouse, rat, dog, C. elegans and Drosophila.
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In genetics, micro RNAs (miRNA) are single-stranded RNA molecules of about 21-23 nucleotides in length which regulate gene expression. In other words, a non-coding RNA molecule of approximately 21-23 nucleotides that inhibits mRNA expression is known as micro RNA.
The main points about micro RNA are given below:
1. Micro RNA is involved in regulation of gene expression.
2. In a cell, miRNA is transcribed from DNA but not translated into proteins.
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3. Micro RNAs are non-coding molecules of approximately 21-23 nucleotides.
4. Micro RNAs inhibit the expression of mRNA molecule.
5. Mature miRNA molecules are partially complementary to one or more messenger RNA (mRNA) molecules.
6. The main function of miRNAs is to down regulate gene expression.
7. It has been reported that a typical mammalian cell contains as many as 50,000 different miRNAs.
8. Micro RNAs were first described nor the worm C. elegans in 1993.
9. The term micro RNA was only introduced in 2001.
10. Only one strand of DNA can function as templates to give rise to miRNA.
Formation of Micro RNAs (miRNA):
The formation of micro RNAs consists of three important steps, viz:
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(i) Formation of primary miRNA,
(ii) Formation of precursor miRNA, and
(iii) Formation of mature functional miRNA.
These are discussed below:
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1. Formation of Primary miRNA:
The primary transcript is synthesized from DNA template. The miRNAs are first transcribed as primary transcripts or pri-miRNA with a cap and poly-A tail and then processed to pre- miRNA. Either the sense strand or antisense strand of DNA can function as templates to give rise to miRNA.
2. Formation of Precursor miRNA from pre-miRNA:
A short, 70-nucleotide stem-loop structure known as pre-miRNA is formed from primary miRNA in the cell nucleus. This processing is performed in animals by a protein complex known as the Microprocessor complex, consisting of the nuclease Drosha and the double-stranded RNA binding protein Pasha.
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3. Formation of Mature miRNA from pre-miRNA:
These pre-miRNAs are then processed to mature miRNAs in the cytoplasm by interaction with the endonuclease Dicer, which also initiates the formation of the RNA-induced silencing complex (RISC). This complex is responsible for the gene silencing observed due to miRNA expression and RNA interference. The genes encoding miRNAs are much longer than the processed mature miRNA molecule.
Thus microRNA (miRNA) is produced from precursor microRNA (pre-miRNA), which is formed from a microRNA primary transcript (pri-miRNA). The process of formation of miRNA can be represented as follows.
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In plants, this pathway differs slightly because plants lack Drosha homologs. In plants, Dicer homologs alone effect several processing steps. The pathway is also different for miRNAs derived from intronic stem-loops. These are also processed by Dicer but not by Drosha.
Processing of Micro RNAs (miRNA):
Efficient processing of pre-miRNA by Drosha requires the presence of extended single-stranded RNA on both 3′- and 5′-ends of hairpin molecule. These ssRNA motifs could be’ of different composition but their length is important for processing. A bioinformatics analysis of human and fly pri-miRNAs revealed very similar structural regions, called ‘basal segments’, ‘lower stems’, ‘upper stems’ and ‘terminal loops’.
Based on these conserved structures, thermodynamic profiles of primiRNA have been determined. The Drosha complex cleaves RNA molecule -2 helical turns away from the terminal loop and ~1 turn away from basal segments. In most analysed molecules this region contains unpaired nucleotides and the free energy of the duplex is relatively high compared to lower and upper stem regions.
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Most pre- miRNAs do not have a perfect double-stranded RNA (dsRNA) structure topped by a terminal loop. Clear similarities between pri-miRNAs encoded in respective (5′- or 3′-) strands have been demonstrated.
The pre-miRNA stem-loop is cleaved by Dicer into two complementary short RNA molecules, but only one is integrated into the RISC complex. This strand is known as the guide strand and is selected by the argonaute protein. The remaining strand, known as the anti-guide or passenger strand, is degraded as a RISC complex substrate.
After integration into the active RISC complex, miRNAs base pair with their complementary mRNA molecules and induce mRNA degradation by argonaute proteins, which are catalytically active members of the RISC complex.
Differences Between Small Interfering RNA (siRNA) and Micro RNA (miRNA):
Micro RNA [miRNA] is a short (about 21 to 23 nucleotides) single-stranded RNA molecule that is now recognized as playing an important role in gene regulation. It has some similarities and some differences with small interfering RNA (siRNA).
Both miRNA and siRNA have gene regulation functions, but there are slight differences. The miRNA may be slightly shorter [21-23 nucleotides] than siRNA (20 to 25 nucleotides). The miRNA is single-stranded, while siRNA is formed from two complementary strands. The two kinds of RNAs regulate genes in slightly different ways.
The miRNA attaches to a piece of messenger RNA (mRNA)—which is the master template for building a protein – in a non-coding part at one end of the molecule. This acts as a signal to prevent translation of the mRNA into a protein. siRNA, on the other hand, attaches to a coding region of mRNA, and so it physically blocks translation.
Roles of Micro RNA [miRNA]:
The miRNAs play important role in gene regulation. Micro RNAs arc also expected to be useful in detection of various diseases and their treatment in the years ahead.
The role of miRNAs in gene regulation and disease detection are briefly discussed as follows:
1. Gene Regulation:
The important cellular function of miRNAs is related to gene regulation. The miRNA is attached to the mRNA at a specific point and inhibits protein translation. In other words, the miRNA complex blocks the protein translation machinery. This is thought to be the primary mode of action of plant miRNAs.
In such cases, the formation of the double-stranded RNA through the binding of the miRNA leads to the degradation of the mRNA transcript. It is also believed’ that miRNA can prevent translation without causing degradation of the mRNA.
2. Micro RNA and Diseases:
The discovery of miRNA has opened up new areas of research. Now miRNA-based diagnostics and therapeutics are getting increasing importance. The miRNA technology is expected to help in diagnosis and treatment of serious diseases like cancer, heart diseases and diseases related to the nervous system.
This will also help in reclassification of different types of cancers. Thus miRNA technology has wide applications in several areas such as cardiac research; virology, cell biology in general and plant biology.
Studies on miRNA expression profiling demonstrated that expression levels of specific miRNAs changed in diseased human hearts, pointing to their involvement in cardiomyopathies.
Furthermore, studies on specific miRNAs in animal models have identified distinct roles for miRNAs both during heart development and under pathological conditions, including the regulation of key factors important for cardio genesis, the hypertrophic growth response, and cardiac conductance. Similarly, several miRNAs have been found to have links with some types of cancer.
