Structure, Function and Utility in Medicine
Structure of RNA
Ribonucleic acid (RNA) is a biopolymer, consisting of individual nucleotides. Like DNA, RNA nucleotides are very similar in that there are four nitrogen bases that make up its sequence. Three of the nitrogenous bases are absolutely identical to DNA: adenine, guanine and cytosine. However, instead of a thymine base as in DNA, there is a structurally close pyrimidine in RNA called uracil. Also, unlike DNA, RNA consists of a single strand, not a duplex. RNA is utilized in many ways in the cell, either as messenger RNA (mRNA), transfer RNA (tRNA) or ribosomal RNA (rRNA). In contrast to the deoxyribose sugars in DNA, ribose sugars that form RNA are less stable and easily degrade. Also, RNA is typically single-stranded, shaped as a hairpin, or double-stranded as in viruses.
In contrast to DNA, RNA is not a single long strand of genetic material that contains all the genes. Each gene is broken up into many introns and exons, with each gene having its own start/stop properties. The exon regions of DNA, which are the parts of sequence that contain useful instructional sequence, are read by RNA polymerases and the resultant exon fragments are pieced together to form a single mRNA transcript whose sequence is completely dependent on the genetic code of the template DNA sequence.
Function of RNA
RNA is transcribed from DNA when a particular gene is needed by the cell. Unlike DNA which is maintained at a constant level (i.e. 23 pairs of chromosomes), the amount of mRNA in a cell is continually changing. Depending on the type of cell and the function of a particular gene in response to a stimuli, the cell may require a massive production of one gene (i.e. upregulation) and zero transcription of another gene (i.e. downregulation).
Messenger RNA transcripts are translocated out of the nucleus and into the cytoplasm where protein synthesis occurs. At this point, tRNA molecules bring amino acids to ribosomes where rRNA links the amino acids together to form full length proteins.
Ribosomal RNA makes up the major share of all cellular RNA (up to 80%). This amount of rRNA demands an intensive transcription of genes which code for its production. The genome compensates for this need by providing copies of the genes that code for rRNA. In eukaryotes, this consists from several hundred (~200 at yeast) to tens of thousands of organized genes. Genes coding rRNAs in a human being are also organized into groups and located in central regions of chromosomes 13, 14, 15, 21 and 22.
RNA is manufactured in the cell during transcription, where RNA polymerases use DNA as a template to produce complementary RNA. Ribosomes in the cell use the RNA as a template to produce proteins, in a process called translation. Three types of RNA are involved in this unique process: messenger RNA (mRNA), transfer RNA (tRNA) and ribosomal RNA (rRNA). In addition to transcription and translation, RNA is also involved in the activation and deactivation of genes.
Utility of RNA in Medicine
RNA enables extensive benefits to the laboratory and to novel therapeutic medicines. Following well established RNA isolation protocols, RNA can be isolated from tissues of study animals or areas of plants, from bacteria, from RNA-containing viruses and from tumors. Upon isolation, the concentration of RNA can be measured since the nucleotide bases of RNA absorbs ultraviolet light at a wave length near 260 nm. The isolated RNA enables researchers to determine individual gene expression levels of a test sample (i.e. treated or tumor sample) relative to a control (i.e. non-treated or normal tissue sample).
After gathering a full understanding of differential gene expression via qRT-PCR or RNA-Seq, scientists then attempt to genetically correct any cellular alterations by intracellular delivery of RNA molecules via transfection in which RNA is reintroduced into cells by electroporation or using liposomes that fuse with the cell membrane to deliver the RNA payload to the interior of the cell. The resultant delivery and resultant downregulation of the target RNA molecules is due to RNA interference (RNAi) in which innate mechanisms find and then regulate or degrade the complementary mRNA sequence of the transfected RNA (i.e. miRNA, siRNA, piRNA). RNAi effects have been induced in both mammalian cells and in tissues for endogenous and alien genes. The resultant loss of expression can be partial or complete, allowing the response to be tailored to the needs of the experiment. A crucial consideration in RNAi products is the length of the double-stranded RNA (dsRNA), as any strand longer than thirty base-pairs may trigger an interferon response leading to cleavage of mRNA and possibly apoptosis. Care must also be taken in the design of RNAi therapeutics such that they do not induce innate immune responses or normal cell functions.
Double-stranded RNA can be delivered in several ways:
- Viral induction https://en.wikipedia.org/wiki/RNA_interference
- siRNA http://en.wikipedia.org/wiki/Small_interfering_RNA
- Plasmid producing shRNA http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2802021/
- miRNA http://en.wikipedia.org/wiki/MicroRNA
RNA has been harnessed for its ability to silence genes or knockdown expression, where a small complementary fragment of RNA is used cellular mechanisms to limit expression of a gene. In vivo transfection (mRNA or siRNA transfection) is known to be difficult due to RNA stability. Some companies have developed specialized RNA transfection reagents (see transfection kits) to help researchers achieve high transfection efficiency both in vitro and in vivo.